Childs Et Al., 2021 ING

The Importance of Leptin to Reproduction
Downloaded from https://academic.oup.com/endo/advance-article/doi/10.1210/endocr/bqaa204/5962079 by San Francisco State University Library user on 14 November 2020
Gwen V. Childs1, Angela K. Odle1, Melanie C. MacNicol2 and Angus M. MacNicol2
Department of Neurobiology and Developmental Sciences,
University of Arkansas for Medical Sciences, Little Rock, AR 72205
1
These first authors contributed equally to the manuscript
2
These authors contributed equally as Senior Authors on this manuscript
t
ip
cr
Correspondence and reprint requests should be sent to Gwen V. Childs, Ph.D.
us
Grants supporting this work: NIH R01 HD059056 (GVC); NIH R01HD087057 (GVC and AMM);
NIH R01HD093461 (AMM, GVC, and MCM), NIH R01DK113776-01 (GVC, AMM, and MCM);
NIGMS P20 GM103425 and P30GM11070 (Dr. Edgar Garcia-Rill)
an
M
The authors have nothing to disclose

e d
pt
ce
Ac
© The Author(s) 2020. Published by Oxford University Press on behalf of the Endocrine
Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com
Abstract
A healthy nutritional state is required for all aspects of reproduction and signaled by the
adipokine leptin. Leptin acts in a relatively narrow concentration range; too much or too little will
compromise fertility. The leptin signal timing is important to prepubertal development in both
sexes. In the brain, leptin acts on ventral premammillary (PMV) neurons which signal kisspeptin
(Kiss1) neurons to stimulate gonadotropin releasing hormone neurons (GnRH). Suppression of
Kiss1 neurons occurs when AgRP neurons are activated by reduced leptin, because leptin normally
t
ip
suppresses these orexigenic neurons. In the pituitary, leptin stimulates production of GnRH
cr
receptors (GnRHR) and follicle-stimulating hormone (FSH) at midcycle, by activating pathways
that de-repress actions of the mRNA translational regulatory protein Musashi. In females, rising
us
estrogen stimulate a rise in serum leptin, which peaks at midcycle, synchronizing with nocturnal
an
Luteinizing hormone (LH) pulses. The normal range of serum leptin levels (10-20 ng/ml) along
with gonadotropins and growth factors, promote ovarian granulosa and theca cell functions and
M
oocyte maturation. In males, the prepubertal rise in leptin promotes testicular development.
However, a decline in leptin levels in prepubertal boys reflects inhibition of leptin secretion by
d
rising androgens. In adult males, leptin levels are 10-50% of those in females, and high leptin
e
inhibits testicular function. The obesity epidemic has elucidated leptin resistance pathways with too
pt
much leptin in either sex leading to infertility. Under conditions of balanced nutrition however, the
ce
secretion of leptin is timed and regulated within a narrow level range that optimizes its trophic
effects.
Ac
Keywords: Leptin, Leptin receptors, Pubertal development, Ventral premammillary nucleus (PMV),
gonadotrope, granulosa and theca cells, oocytes, Leydig cells, Sertoli cells, spermatocytes.
2
Introduction
The Impact of Nutrition on Reproduction. Food availability is considered the single most
important environmental factor that influences mammalian reproduction (1). Healthy reproduction
requires adequate nutrition for appropriately timed maturation events and the development of
secondary sex characteristics (2,3). Maternal nutrition is required for the timing of normal
reproductive cyclicity to promote the development of healthy female gametes, and prepare the
female to support a pregnancy and provide milk for the young (4,5). Adequate nutrition is also
t
ip
important for female receptivity to mating, the timing of which ensures that the short-lived sperm
cr
can reach a viable oocyte (6).
us
Nutritional deficiency results in reduced pituitary Luteinizing hormone (LH) release in response
to decreased hypothalamic Gonadotropin-releasing hormone (GnRH) stimulation (7-10). When

an
reproduction occurs in spite of nutritional deprivation, the initial outcomes are a reduction in the
number and/or the size of the young (2,3). Severe nutritional deficiency inhibits reproduction
M
altogether as the immediate survival of the animal takes priority over its reproduction (2,3).
d
In order to respond to nutritional challenges, metabolic signals relay information on energy

e
status to the Hypothalamic-Pituitary-Gonadal (HPG) axis. One of the most powerful signals is
pt
leptin, the 167 amino acid product of the Lep (formerly ob) gene. Leptin has the distinction of being
the only known biomarker of adiposity, as its circulating levels are in linear proportion to fat mass
ce
(11,12).
Ac
The first recognized role for leptin was to send a negative feedback signal to the brain when energy
stores were adequate, resulting in decreased food intake and increased energy expenditure (11,13-16).
However, studies of animals that are leptin-deficient(10,17-19), revealed that leptin also plays a
dynamic role in reproduction (17-27).
History of the discovery of leptin. Researchers sought answers to questions related to
nutritional state and reproduction long before leptin was discovered(28), as it has been recognized
that an animal must grow to maturity and attain a threshold of body fat before it can breed(2). Clues
3
to a factor that signaled the amount of adiposity were uncovered 70 years ago in a mouse line
bearing spontaneous gene mutations that rendered it obese (ob/ob)(29) or diabetic (db/db)(30).
Studies of these obese mouse lines also reported that females and most males were infertile because
their HPG axis was immature(31).
Classical parabiosis experiments by DL Coleman showed that the db/db mice carried a blood-
borne factor that caused significant weight loss and starvation in its parabiotic partner in pioneering
work which led to the Lasker prize(28,32). Coleman hypothesized that the ob/ob mouse lacked the
t
ip
secreted factor, and the db/db mouse overproduced it, but could not respond to it (28,32,33).
Coleman's group reported that the factor was a component of adipose tissue but were never able to
cr
isolate it. In 1994, Zhang et al (in Dr. Friedman's laboratory) successfully isolated the factor by
us
positional cloning, naming it leptin after the Greek "leptos" meaning thin(13). The ob/ob mice
carried a mutation in the leptin gene (Ob or Lep) and db/db mice had a mutated leptin receptor gene
an
(Db or Lepr)(34-37).
M
Regulation of leptin secretion from adipocytes. As reviewed by Cammisotto et al (38), studies
of humans and rodents since its discovery have shown that leptin transcription and secretion in
d
adipocytes may be stimulated by dexamethasone and other glucocorticoids and peroxisome

e
proliferator-activated receptor-gamma agonists. Leptin is inhibited by catecholamines, free fatty

pt
acids and thyroid hormones. Studies of rats reported that both glucose and insulin are stimulatory
ce
(39).
Cammisotto et al (38) also reported that leptin secretion from male rat adipocytes was stimulated
Ac
by nutrient signals, including glycolytic substrates (glucose, fructose or pyruvate). In addition,
amino acids L-glutamate stimulated leptin alone; L-aspartate, L-valine, L-methionine and L-
phenylalanine potentiated glucose action and L-Leucine was stimulatory only in the presence of
glucose.
In adult human male or female adipose tissue (40), pituitary thyroid stimulating hormone (TSH)
stimulated leptin by directly regulating adipocytes. However, this same group reported no
4
stimulation by prolactin, Adrenocorticotropin (ACTH), follicle stimulating hormone (FSH) or
luteinizing hormone (LH). As part of its lipolytic function, however growth hormone (GH) reduced
leptin gene expression and leptin secretion in bovine or rat adipocytes (41,42).
With regard to regulation by reproductive hormones, rising estradiol levels are correlated with
rising serum leptin in human and rodent females and estrogen treatment stimulates serum leptin
(43,44). In contrast, androgens inhibit leptin expression (45) and testosterone treatment reduced
leptin levels in adult men (46). Similarly, studies of 3T3-L1 adipocyte tumor cells treated with
t
ip
dihydrotestosterone showed an 80% reduction in Lep mRNA while estradiol increased Lep mRNA
by 140% (46).
cr
This minireview will describe the current state of our knowledge regarding the importance of
us
leptin to reproduction. Its importance was recognized historically in classical studies of ob/ob
an
mice, in which exogenous leptin restored reproduction before it cured the obesity (10,17-19).
However, the mechanism by which leptin influences reproduction is only now becoming clear.
M
Questions remain including whether leptin regulates the timing of events in the reproductive system
under normal energy conditions and whether it is involved in the timing of the reproductive cycle.
d
We will address the target tissues and cells involved and discuss how leptin influences each.
e
pt
Control of Leptin Signaling

ce
Pleiotropic Leptin Receptors. The leptin receptor (LEPR) (47,48), a product of the Lepr gene
Ac
and a member of the class I cytokine receptor superfamily has 6 isoforms, which may be expressed
in different proportions depending on the cell type and the species(47-53). LEPRa,b,c,d,f have
identical extracellular and transmembrane domains but differ in the length of their intracellular tail.
Short isoforms (LEPRa, c, d, f) have 30-40 residue intracellular tails and unique C-termini,
suggesting unique roles(47). LEPRa is broadly expressed and may facilitate leptin transfer across
5
the blood-brain barrier(47). The sixth isoform, LEPRe is soluble, secreted into the bloodstream and
may transport leptin, modulating its bioavailability(47).
LEPR forms dimers or oligomers and can heterodimerize in the presence and absence of
leptin (47). LEPRs are distributed intracellularly in target cells in the receptor-mediated endocytic
pathway, with only 10-20% of receptors found on the extracellular surface and remaining molecules
in the endoplasmic reticulum, trans-Golgi, and endosomes, available for recycling to the plasma
membrane as needed(48).
t
ip
LEPR signaling pathways. The best characterized signaling pathway activated by the long
cr
isoform LEPRb, is the Janus kinase (JAK) and Signal transducer and activator of transcription
proteins [STAT3 and STAT5(48)]. Paradoxically, STAT5 has been implicated in reproductive
us
competence (54,55). However, ablation of STAT5 and/or STAT3 in cells expressing LEPR (56)
an
results in normal puberty onset, cyclicity and fertility over a 4-month period, suggesting that HPG
target cells process leptin signals through multiple signaling pathways. These leptin-signaling
M
pathways involve Mitogen-activated protein kinase/Extracellular signal regulated kinase (ERK) 1/2
(MAPK) (57,58); Phosphoinositol 3 kinase (PI3K) (59,60), mammalian Target of rapamycin

d
(mTOR)(61-63) and/or nitric oxide (64). Investigators studying the individual target cells in the
e
HPG axis have included tests of each of these pathways to determine which are involved in leptin
pt
actions. Some of these studies will be discussed in later sections focused on the leptin target cells.
ce
Leptin resistance. As leptin operates in a narrow concentration range(65), elevated leptin levels,
such as those seen in obesity, are often accompanied by resistance to leptin signaling, (66). This
Ac
multifactorial event largely centers on the trafficking and signaling of LEPR (47,48,67) and
involves feedback inhibition via Suppressor of cytokine signaling 3 (SOCS3), protein tyrosine
phosphatase 1B (PTP1B), and T-cell PTP (TCPTP) (47,48,67). The expression of these negative
regulators is elevated in the hypothalamus of obese animals. Deletion of SOCS3 leads to enhanced
leptin actions and attenuation of leptin resistance in models of diet-induced obesity(47,48).
6
Circulating leptin: which is more important, levels or timing?
Proof that adipocytes are the source of circulating leptin. Many reviews have identified
changes in serum leptin levels critical to metabolic health, puberty, and the reproductive cycle (17-
27). However, in addition to adipocytes, leptin production has been detected in numerous organs
leading researchers to speculate that each of the organs may contribute to differential circulating
leptin [reviewed in(68)]. Variation in leptin levels may reflect differential secretion from these
organs, or differential regulation of adipocytes. At this point, no systemic functions for extra-
t
ip
adipocyte leptin have been reported. Our laboratory addressed this with tissue-specific Lep-null
cr
mouse models that selectively ablated leptin in adipocytes or pituitary
us
somatotropes(68). The ablation of adipocyte leptin resulted in undetectable serum leptin levels in
adult or neonatal mice (Figure 1A), because it caused obesity similar to that of ob/ob mice. In
an
contrast, mice lacking leptin only in somatotropes did not show a decrease in serum leptin (Figure
1A). These findings point to adipocytes as the main, if not the only source of circulating leptin(68)
M
and are consistent with the tight association between circulating leptin and adiposity.
d
The neonatal leptin surge, puberty and metabolic health. The association between the pre-
e
pubertal increase in fat stores and the onset of puberty originally supported a hypothesis that the leptin
pt
signal is vital for the timing of puberty. Reports that leptin accelerates puberty (16,69,70) suggested
that leptin might be the primary metabolic trigger. However, there was no correlation between
ce
prepubertal serum leptin levels and the timing of puberty in normal rodents (71-73) or primates (74-
Ac
77) arguing against this hypothesis. There is evidence, however, for a rise in leptin during the third
trimester in the human fetus (78,79) or postnatally in rodents (72,73,80,81), shown in Figure 1B.
The data in Figure 1B are the first to compare postnatal male and female mice, showing a
postnatal age variance, but no sex differences (Two-way ANOVA followed by Tukey's posthoc test.
See Figure legend). These studies are important because previous studies reported data only from
female mice (80), male rats (82), or data from rats (83) or mice (84) which combined the sexes.
Studies show that this neonatal leptin rise does not coincide with an increase in fat mass and is
7
unrelated to appetite regulation. Indeed, during this period of development, leptin does not inhibit
food intake(85,86). However, the period of the rise is critical, because, in leptin-deficient mice, leptin
restored maturation and development only if given during the neonatal period (87,88). The
importance of the neonatal leptin rise to reproduction and metabolism was highlighted recently in a
study by Ramos-Lobo et al (89), who studied an inducible mouse model which were LEPR-null at
birth and then were rescued globally during the fourth or 10th week of age (89). Whereas some
systems recovered, the reproductive system remained immature or dysfunctional in most animals.
t
Males retained low testicular weights and exhibited low GnRH fiber density and kisspeptin (Kiss1)
ip
mRNA. Only 10% of the wild type females mated with rescued males became pregnant. However,
cr
50% of rescued females became pregnant in spite of dysfunctional cycling as defined by vaginal
us
smears. Collectively the study pointed to the importance of leptin signals early in development for
metabolic and reproductive function in the adult.

an
Studies that manipulated the neonatal leptin surge have reported that it can be blunted or altered
by maternal undernutrition (82,84) or administration of a leptin antagonist(90), with diverse

M
consequences, including the improper formation of hypothalamic networks involved in feeding and
d
dysfunctional metabolic responses (82,91,92). Maternal undernutrition blunted or shifted the neonatal
e
leptin surge, with either result having lifelong detrimental effects on the metabolic health of the
pt
offspring (82,84,91,92). Progeny exhibit sensitivity to a high fat diet (HFD)(83). Another study (84)
replicated the conditions of an early leptin surge by injecting leptin on postnatal day 5, resulting in
ce
high sensitivity to a HFD in the treated adults. This study thus introduced the concept that the timing
Ac
of the leptin rise may be as important as serum levels for early developmental processes.
Researchers investigating effects of altering the leptin surge on the reproductive system found that
blockade or alteration of the postnatal leptin surge decreased testicular growth and FSH in male rat
pups and decreased ovarian growth and FSH in females (93). Puberty onset was delayed in both sexes
(93) and ovarian primordial follicles were reduced in females (90). There were sex-dependent effects
on the development of the hypothalamic circuits that regulate reproduction (especially the kisspeptin
network) (94-96).
8
Our recent studies of mice lacking LEPR in gonadotropes have demonstrated a leptin dependency
for expression of GnRH receptor proteins, Fshb mRNA, and the mRNAs for the FSH regulator activin
(97-99), which may have begun during neonatal development in females. Female mice lacking LEPR
in gonadotropes have significantly reduced levels of GnRHR, and Fshb, Actßa and Actßb mRNAs
(97-99). We propose that this dependency may be manifested as early as neonatal development,
because studies of developing purified gonadotropes by Wen et al reported that full co-expression of
GnRHR and FSH coincides with the rise in serum leptin (by postnatal day 7) (100). Additional
t
studies of female rat pituitaries reported a rise in Fshb mRNA during the time of the postnatal rise in
ip
leptin (101-103), along with a parallel rise in pituitary activin ßa and ßb subunits (104).
cr
We have recently performed a maternal undernutrition study on mice with a 20% food restricted
us
diet. The underfed progeny had an early leptin surge, and female mice showed delayed puberty, in
spite of the fact that they had sufficient leptin at the time of the normal surge. On day 29, 90.1% of
an
11 controls and only 12.5% of the 8 underfed progeny showed vaginal opening. The remaining
underfed progeny did not show vaginal opening until day 32 (Miles et al, Ms in preparation). These
M
ongoing studies continue to indicate that the timing of the neonatal leptin surge is as important to
reproductive and metabolic health as the level.

e d
The Importance of Leptin Pulses to the Timing of Puberty. The timing of the leptin signal was
pt
initially implicated as a positive regulator of puberty, because injections of leptin in weanling
female mice caused early onset of puberty(16). In male monkeys, when the time of puberty onset is
ce
defined by the nocturnal LH pulses, studies showed that nocturnal leptin levels increased
Ac
significantly before puberty onset along with a gradual increase in Growth hormone (GH) and
Insulin-like growth factor-1 (IGF-1) levels (105). The investigators suggested that leptin stimulates
the GH-IGF-1 axis, which in turn stimulates GnRH and LH. Nocturnal leptin secretion was also
increased by 28 days of pubertal development in female rats (106). Thus, studies of leptin secretion
during postnatal and peripubertal development point to the importance of both nocturnal timing and
amplitude as mechanisms leptin uses to permit development of adult reproductive competence.
9
Sex differences in Leptin Secretion in the Adult. Figure 1B shows identical leptin secretory
patterns in postnatal male and female mice. However, in adult humans there are striking sex
differences in serum leptin concentrations beginning in the peripubertal period (107). In humans,
this begins as early as 10-years of age in boys, which show a decline in serum leptin during puberty.
Normal weight (BMI <30) human women have leptin levels averaging 23.5 ± 1.5 ng/mL with a
range between 4.7-46 ng/mL(108). In contrast, healthy weight human men average 9 ± 0.83 ng/mL
serum leptin with a range of 2.65-20.7 ng/mL. Indeed, the average leptin levels in women are close
t
to obesity levels for men (29 ± 1.55 ng/mL). This difference is likely due to sex steroids as
ip
estrogens stimulate leptin release by adipocytes, whereas androgens inhibit leptin release [reviewed
cr
in (109)] . This sets up a negative association between leptin and testosterone levels in males, which
us
reflects androgen's inhibitory influence upon adipocyte leptin (46,110) and explains the peripubertal
reduction in serum leptin in males (109).

an
Cyclicity of leptin secretion in the female. Not only is leptin secretion higher in the human
female, it exhibits cyclicity, which was first reported as an increase from 14.9 ng/ml in the early
M
follicular phase to 20.4 ng/ml at the mid-luteal phase (65). Further studies also reported synchrony
d
between nocturnal leptin and LH pulses in normal cycling women(111,112). This synchrony with
e
leptin, LH and estradiol pulses was best seen at night, supporting a hypothesis that leptin regulates
pt
LH and estradiol oscillations. Many additional studies of human females showed cyclic changes in
serum leptin as cited in ref (113), which reports a comprehensive study of 259 healthy
ce
premenopausal women (509 cycles) documenting hormonal changes during both ovulatory and
Ac
anovulatory cycles (113). They assayed serum leptin, estradiol, progesterone, LH, FSH and
testosterone 8X per cycle and factored exercise and body mass index (BMI) into their analysis.
They reported an increase in serum leptin from 16.7 ng/ml to 20.4 ng/ml in the luteal phase with
leptin levels averaging 21.7 ng/ml at the time of the LH surge (113). The highest levels of leptin were
associated with higher estradiol, progesterone, ovulatory LH, testosterone, but lower FSH. It is
unclear why high levels of leptin were not correlated with high FSH levels. The explanation may be
that the best correlation between leptin levels and gonadotropins is evident following nocturnal assays
10
of leptin and LH pulses (111,112). It is significant however that overall leptin levels were lower in
anovulatory cycles, which pointed to the need for normal development of ovarian follicles to produce
the estrogen. This study suggested that the source of the rising leptin might be from ovarian follicles
(113). However, our recent studies showing undetectable serum leptin levels in mice lacking leptin
only in adipocytes argue against an ovarian contribution to serum leptin (68) (Figure 1A).
Leptin Interactions with Hypothalamic-Pituitary-Gonadal Target Cells
t
Leptin target cells in the hypothalamus. The search for leptin-regulatory pathways in the brain
ip
was begun in pioneering studies(114) which partially ablated exon 17 of LEPR (thus eliminating the
cr
signaling domain of LEPRb) in hypothalamic neurons. They reported fertility with a partial ablation,
us
but infertility only after LEPRb was ablated in all target neurons. The search then began for the subset
of LEPR neurons specifically involved in fertility. This search was challenging because GnRH
an
neurons lacked LEPR (115,116). Moreover, as reviewed by Elias and Purohit, LEPR expression in
Kisspeptin neurons was low or varied with the species or physiological state(22). Furthermore,
M
ablation of LEPR in Kiss1 neurons had no effect on reproduction (117). This field has been
reviewed comprehensively, and the reader is referred to a series of excellent papers for further
d
details on early work in this area (1,19-23,25,118-120). This minireview will concentrate mainly on
e
recent findings focusing on two major sets of leptin-responsive neurons that mediate either
pt
stimulation or suppression of the reproductive system.

ce
Donato et al (117,121) discovered that neurons in the ventral pre-mammillary nucleus (PMV)
Ac
formed a pathway mediating leptin stimulation of Kisspeptin neurons, which then stimulated GnRH
neurons (Figure 2). This was initially based on studies in rats showing that PMV lesions blunted
kisspeptin and GnRH neuronal activity during the afternoon of proestrus, decreasing LH and
estrogen secretion and disrupting estrous cyclicity. As stated above, this group had also determined
that deletion of LEPR in Kiss1 neurons did not affect the timing of puberty or fertility (117), ruling
out Kiss1 neurons themselves as direct leptin mediators of reproduction. They then developed a
mouse model, which re-expressed LEPR in the PMV of a LEPR-knockout mouse model and
11
reported that restoration of LEPR in the PMV induced puberty, improved estrous cycles and
allowed pregnancy (117,121). However, only a fraction of the females (20%) carried pups to term.
A follow-up study (122) correlated restoration of a leaner weight with the delivery of healthy pups
in 1/5 females. In a different model in which Cre-recombinase was delivered to the PMV by an
adenovirus(122), puberty was restored in all 11 mice which had correctly targeted the PMV (out of
18 tested). Five of these mice became pregnant, and 1 of these had a healthy litter. Thus,
collectively, these studies demonstrated the importance of the PMV neurons as leptin targets for
t
stimulation of Kisspeptin neurons in the anteroventral periventricular and caudal arcuate nuclei and
ip
regulation of pubertal development, cyclicity and pregnancy (117,121,122). The low pregnancy
cr
success rate following selective restoration of LEPR may have reflected the mouse obesity and the
us
lack of LEPR on extrahypothalamic target cells.
The investigations of stimulatory pathways then continued with studies to determine the identity
an
of the neurotransmitters in PMV neurons responsible for leptin-mediated regulation of either GnRH
neurons or Kisspeptin neurons. As shown in Figure 2, three candidates regulators are glutamate
M
(22,120,123), pituitary adenylate cyclase activating polypeptide (PACAP) (123) and nitric oxide
d
(64,120,124). PACAP regulates both food intake and gonadotropin release (123). Ross et al (123)
e
reported that PACAP was expressed in LEPR-bearing PMV neurons that produce glutamate and
pt
had the highest levels of pSTAT3 activity, an indicator of LEPR function. They selectively ablated
PACAP in all LEPR-expressing neurons and reported that females showed delayed onset of
ce
puberty, a blunted LH surge, dysregulated estrous cycling and low numbers of pups/litter (123).
Ac
Tests of responses to exogenous leptin showed a blunted LH response. However, exogenous
kisspeptin given to these PMV neuron PACAP-null mice resulted in stimulated LH secretion
suggesting that GnRH neurons were intact and that the dysfunction in the connection was at the
level of the kisspeptin neurons. When PACAP was deleted selectively in adult animals (with
adenovirus carrying Cre-recombinase) females had longer cycle lengths, fewer cycles in a 25-day
period, reduced corpora lutea, delays in the time to pregnancy and a failure to nurture their pups.
Tests of LH responses to exogenous kisspeptin showed intact signaling between kisspeptin and
12
GnRH neurons. However, exogenous leptin stimulated higher levels of LH. This aspect of the
study suggested that PACAP from the PMV may be needed for normal Kiss1 to GnRH neuronal
relays but perhaps not for the relay of the leptin signal. Nevertheless, their studies mapped distinct
connections from the leptin receptive PMV PACAP neurons to both sets of Kisspeptin neurons and
they showed PACAP stimulation of a subset of these neurons(123).
As investigators were identifying pathways mediating the stimulation of Kiss1 and GnRH,
parallel studies investigated pathways that may mediate the suppression of GnRH secretion during
t
ip
times when leptin signals are low or absent, such as during food deprivation. Candidate pathways
involved GABAergic leptin receptor-bearing neurons (118) based on findings showing that deletion
cr
of LEPR specifically from GABAergic, but not Glutaminergic neurons caused infertility in females
us
(125,126). Zuure et al (125) reported that deletion of LEPR in GABA neurons in mice did not
interfere with the estrogen negative feedback to GnRH neurons. However, the ablation of leptin
an
signals clearly blunted a preovulatory-like LH surge. In their comprehensive discussion of possible
sites for these GABAergic neurons (125), they pointed to studies of agouti-related peptide (AgRP)
M
neurons in the arcuate nucleus, which also produce neuropeptide Y (NPY). These are illustrated in
d
Figure 2. Zuure et al hypothesized that these factors may serve as intermediates that suppress HPG
e
neuronal secretion when leptin signaling is reduced (in fasting, for example). This hypothesis was
pt
based on the fact that normally leptin inhibits this group of orexigenic neurons, so the lack of leptin
signals would result in increased AgRP and NPY. Zurre et al cited evidence for AgRP and NPY
ce
inhibitory influences on GnRH and Kiss1 neurons(125). In addition, NPY and AgRP are elevated
Ac
in mice lacking leptin (ob/ob mice) (127,128).
Early evidence for the involvement of AgRP neurons in reproduction was strengthened when
Wu et al (128) reported that ablation of AgRP neurons in adult ob/ob mice restored fertility. Five
years later, Egan et al (129) selectively deleted leptin receptors in AgRP neurons and reported
delays in puberty in female but not male mice. However, reproduction was not prevented. When
LEPR was restored in AgRP neurons in infertile Lepr-null mice, they reported restoration of all
reproductive attributes, although puberty was delayed in females. The restored animals remained
13
subfertile, however, and females were prone to dystocia complications because of their obesity.
They did show a normal LH response to removal of estrogen feedback indicating that the Kiss1-
GnRH circuit was intact in the rescued animals.
Most recently, the AgRP pathways were further delineated in mice by Padilla et al(127) as they
stimulated AgRP fibers and detected inhibitory synaptic connections with kisspeptin neurons in
both Arcuate and AVPV neurons, which were reduced when AgRP neurons were ablated (Figure
2). In addition, when the AgRP neurons were chemogenetically activated, the females showed
t
ip
delayed estrous cycle length and decreased fertility. Their tests of GABA responses by GnRH
neurons indicated that the AgRP neurons mediate their suppression of fertility by directly inhibiting
cr
Kiss1 neurons.
us
To summarize, the studies thus far indicate that leptin-dependent PMV neurons synapse on
an
Kiss1 neurons and their signaling with PACAP, glutamate or NO as neurotransmitters is required
for reproduction and the onset of puberty. In addition, reduced leptin signals in states of low energy
M
stores or food deprivation permit arcuate orexigenic neurons to produce AgRP, NPY and GABA to
suppress reproduction and thus conserve energy for food gathering. The next sections of this
d
minireview will focus on the downstream target cells in the HPG axis to present evidence for leptin
e
integration with HPG regulatory hormones to amplify or limit its metabolic signal to the system.
pt
Pituitary Gonadotropes as Leptin Target cells. Many early studies have shown that pituitary
ce
gonadotropes bear leptin receptors and their expression of this receptor varied with the stage of the
cycle. Lepr mRNA was first detected in rat pituitary cells by Zamerano et al (53) and LEPRa and
Ac
LEPRb expression by gonadotropes was first reported by Jin et al (130,131). Evidence supporting
gonadotropes as leptin target cells also came from studies showing that LEPR were functional
(97,130-137). In addition, their leptin dependency was evident by the reduced numbers of
gonadotropes in leptin-deficient mice (135,138-140).
Our cytochemical studies of mice and rats showed that pituitary LEPRb immunolabeling increased
during the luteal phase (97,141), which correlated with the midcycle rise in serum leptin (43,44,142-
14
144). This increase in LEPRb cells reflected additional cells that were dual labeled for LH-beta and
LEPR (97).
Multiple in vivo studies have reported leptin modulation of the expression and/or secretion of
gonadotropins in vivo (31,97,115,143,145-149). For example, leptin restored LH secretion in fasted
mice, rats, and monkeys (144,150-157) and normalized reproductive hormone levels in a leptin-
deficient prepubertal child (158), as well as in adult men (159) and women with amenorrhea
(160,161).
t
ip
In vitro studies have also demonstrated that leptin stimulates LH secretion (64,143,145,149,162).
cr
Our laboratory reported that in vitro exposure to 10-100 pg/mL leptin restored the numbers of
gonadotropes reduced by 24 h of fasting (163). Leptin also stimulated FSH secretion from monolayer
us
pituitary cultures derived from two groups of female monkeys (164). The leptin-mediated pathway to
an
pituitary gonadotropes is illustrated in Figure 3. Our studies of mice lacking LEPR in gonadotropes
indicate that leptin may be involved directly or indirectly in the translation of GnRHR and FSH
M
proteins, in partnership with estradiol positive feedback and pulses of GnRH (97).
Whereas there is agreement about gonadotrope responses to leptin, the importance of gonadotropes
d
as direct metabolic sensors has been a subject of controversy. Are gonadotropes

e
pt
passive mediators of hypothalamic or gonadal responses to metabolic changes? Does leptin play an
active trophic role in the regulation and differentiation of the gonadotrope population? We addressed
ce
these questions using a deletion mutant mouse model in which exon 17 of Leprb (which encodes the
Ac
STAT3 binding site) was ablated selectively in gonadotropes (97). This loss of LEPR signaling in
gonadotropes did not affect the timing of puberty nor did it affect ovarian, uterine or testicular weights
(97). However, mutant females showed delayed time to first pregnancy and lower numbers of pups.
The remaining pups survived indicating normal lactation. With respect to pituitary hormone
expression, mutant diestrous females had reduced serum LH and FSH and reduced mRNA levels of
Fshb and activin. Mutant males had reduced levels of Fshb and Cga mRNAs (97).
15
More recently, we ablated exon 1 in LEPR in gonadotropes (98), which removes the signal peptide
common to all LEPR isoforms, thus preventing the LEPR molecules getting translocated to the ER or
targeted to the cell membrane. Comparing the phenotype of our two lines will be useful as Lepr exon
17-null gonadotropes may still express short forms of LEPRa and signal through the MAPK or nitric
oxide pathways (64,149). Our ongoing studies indicate that gonadotrope Lepr exon 1-null mice are
more severely affected by loss of LEPR in gonadotropes, including more severe subfertility or
infertility and abnormal estrous cycling (98).
t
ip
The most striking change in LEPR-null gonadotropes has been the reduced expression of GnRH
receptors (GnRHR) (97) as detected by biotinylated GnRH or immunolabeling for GnRHR, especially
cr
in females. Furthermore, leptin stimulated GnRHR proteins but did not change mRNA levels (99).
us
These data indicated that GnRHRs were an important post-transcriptional leptin target.
The gonadotrope Lepr exon 17-null mice also exhibited some additional surprising findings,
an
suggesting that leptin's actions on gonadotropes have a broader impact, specifically on cells belonging
to the Pou1f1/Pit1 transcription factor expressing lineage. Both males and females showed reduced
M
serum growth hormone (GH) and males showed reduced serum Thyroid stimulating hormone (TSHb)
d
along with reduced Gh and GH releasing hormone receptor (Ghrhr) mRNA levels. Figure 3 shows
e
the leptin influence on somatotropes as well.

pt
There are several possible explanations for these off-target effects reviewed recently by us (165).
ce
First, we have shown that Pou1f1/Pit1 is another post-transcriptional leptin target (166,167). It is
possible that the targeting construct used to delete gonadotrope LEPR may also function in a
Ac
multipotential Pou1f1/Pit1 pituitary precursor cell population that perhaps become
somatogonadotropes (165,168-172), as they are stimulated by estradiol (173) and following their
expression of GnRHR(170) and GHRHR(171). Alternatively, deletion of LEPR may have affected
the production of paracrine factors secreted by gonadotropes, which stimulate somatotropes and
thyrotropes (174). Further studies are needed to explore roles for leptin in transdifferentiation of
pituitary cells, as well as modulating paracrine interactions in the pituitary. Figure 3 shows that leptin
stimulates gene expression in somatotropes, and more details are reported elsewhere (166,167,175).
16
Other investigators have also recently addressed the relative importance of gonadotrope LEPR
by restoring it in mice genetically engineered to be ubiquitously deficient in LEPR (176). They
reported that although FSH levels were elevated, fertility was not restored (176). This finding is
expected, because the hypothalamic neuronal target cells remained LEPR-null and the mice remained
morbidly obese. Without GnRH from the hypothalamus, gonadotropes will remain in a pre-pubertal
state. Whereas GnRHR was not assayed (176), their findings showing elevated FSH levels confirming
FSH as a downstream leptin target (97).
t
Leptin's actions on gonadotropes: A role for Musashi. After we discovered that leptin regulated
ip
GnRHR through post-transcriptional pathways (97,98), our in silico analysis showed that the 3'
cr
untranslated region (3’UTR) of murine Gnrhr mRNA contains 3 consensus binding sequences for the
us
translational regulatory protein Musashi [MSI, MSI binding elements (MBEs)]. The Musashi1
isoform (MSI1) and Musashi2 isoform (MSI2) are sequence-specific RNA binding proteins that
an
repress target mRNA translation. These proteins promote stem and progenitor cells self-renewal, as
they oppose translation of mRNAs that encode pro-differentiation factors and inhibitors of cell cycle
M
progression (177). Although MSI was reportedly expressed in the mouse pituitary (178), no
functional role had been determined.

e d
We reported that MSI binds to the Gnrhr mRNA 3' UTR and represses translation of a reporter
pt
mRNA driven by this UTR (99). Notably, MSI is a target of regulation by leptin signaling where
leptin stimulation results in decreased Msi mRNA and MSI1 protein levels in GnRHR-cells detected
ce
by immunolabeling and enzyme immunoassay(99). Our findings suggest that Musashi functions to
Ac
regulate differentiated gonadotropes by mediating their expression of GnRHR. Collectively, these
data support the hypothesis that leptin functions to stimulate hormone expression in the gonadotrope,
through its up regulation of GnRHR, activin, and Fshb mRNA levels. We propose that when nutrition
is optimal, the midcycle rise in serum leptin would be permissive for the translation of Gnrhr mRNA
resulting in an activated gonadotrope population(98).
17
Biphasic Actions of Leptin on Target Cells in the Ovary. Early studies showed that ovaries
were among the organs containing the highest levels of Lepr mRNA (179), which stimulated
investigations to determine leptin's direct effects on the ovary. However, following an early report
suggesting that both lean and obese women with polycystic ovarian syndrome had 30% higher
leptin levels (180), researchers hypothesized that leptin was inhibitory (181). Subsequent studies
did not confirm this early study and in fact found that leptin levels were high only in obese PCOS
women, correlating with adiposity (182-185). Unfortunately, however the earliest in vitro studies
t
of human (186,187), rodent (188,189) or bovine (181) (190) granulosa or theca cells used obesity
ip
levels of leptin (>30 ng/ml). These high leptin levels inhibited steroid secretion stimulated by
cr
gonadotropins added with a growth factor (either IGF-1, Transforming Growth factor-alpha) or
us
insulin. High leptin levels also inhibited mouse follicular growth stimulated by FSH, with and
without GH, IGF-1, 8 Br-cAMP, or forskolin (191) and lowered numbers of ovulated oocytes in rats
an
(192). In humans, 50-200 ng/ml leptin also decreased follicles and reduced oocyte maturation(193).
By 2000, the early consensus was that leptin inhibited the synergistic responses to gonadotropins
M
and a growth factor (187-190,194). This conclusion was based on exposure to obesity levels of
leptin. Whereas this supported the hypothesis that obesity impaired ovarian functions (187-
d
190,192), none of the early studies tested the relatively low serum levels of leptin (10-20 ng/ml)
e
found in normal weight women, in spite of the fact that these data were contemporaneously
pt
available(65).
ce
More recent studies utilized in vivo and in vitro models with a full dose range of leptin levels
and discovered leptin's biphasic effects on ovarian target cells. Figure 4 illustrates leptin stimulated
Ac
pathways which influenced follicular development (from primary to antral follicles) by stimulating
theca and granulosa cells in partnership with FSH, IGF-1 and GH. Leptin also stimulated ovulation
and oocyte maturation in partnership with LH. Figure 4 summarizes results from studies in a
number of species, which are reviewed in the following sections. One study showed that induction
of early puberty in rats resulted in granulosa cell hypertrophy, a 2-fold increase in ovulated oocytes
and increases in serum steroid and gonadotropins and ovarian steroidogenic acute regulatory protein
(StAR) and adrenotoxin (ADX) levels (195). In porcine granulosa cells, 10 ng/ml leptin stimulated
18
but 1000 ng/ml inhibited progesterone accumulation and phosphorylation of STAT3 (196).
Similarly, in mice, lower concentrations of leptin increased LH or FSH-stimulated ovarian steroids
and high concentrations decreased follicular growth (197). Leptin stimulated 36% of rat oocytes to
resume meiosis compared with 83% in the presence of LH and FSH (198). Leptin also stimulated
meiotic progression of bovine oocytes at 12.5 ng/ml but inhibited progression at 100 ng/ml (199).
Studies of mechanisms underlying leptin actions in rat ovaries showed similar biphasic effects in
the regulation of Lepr (200), the production of progesterone, and stimulation of ovarian STAT3 and
t
MAPK (at 3-10 ng/ml) (201). The latter study indicated that ovarian cells use short forms of the
ip
leptin receptor. Their in vivo rat ovulation model showed that leptin enhanced ovulation, increasing
cr
phosphorylated STAT3, MAPK and decreasing inhibitory SOCS3 protein, effects that were
us
inhibited by inhibitors of MAPK or JAK/STAT signaling pathways (201). In contrast, an acute high
dose treatment model resulted in reduced ovulation and activated MAPK and STAT3 along with
an
increased inhibitory SOCS3 protein, pointing to mechanisms underlying the development of leptin
resistance as a result of exposure to high concentrations.

M
More examples of leptin biphasic actions were reported in multiple species. In humans, low
leptin (1 and 10 ng/ml) stimulated estradiol and progesterone secretion from granulosa cells, while
d
higher concentrations were suppressive (202). In mice, 10 ng/ml but not 100 ng/ml leptin stimulated
e
germinal vesicle breakdown and appearance of the first polar body (203). In luteinized mouse ovaries,
pt
LH stimulated mRNA levels of the short form LEPRa (Lepra) but not the long form LEPRb (Leprb),
ce
which supported the hypothesis that these cells may use LEPRa (203). In rabbit cumulus-oocyte
complexes, physiological levels stimulated an increase in metaphase II oocytes, actions that were
Ac
impaired with inhibitors of either the JAK/STAT or MAPK pathways (202).
Considering the dynamic changes in the reproductive system, both inhibitory and stimulatory roles
may be physiologically relevant when viewed in the context of the changing serum levels of leptin
during the cycle and with obesity. As described in an earlier section, the nadir point for leptin
secretion in the early follicular phase is 14-16 ng/ml, which rises to reach 21.7 ng/ml at midcycle
(113). Leptin's stimulatory actions described during the last 20 years are evident in those studies that
used that concentration range. It is clear that the HPG system is not geared to respond positively to
19
obesity levels of leptin in the microgram range let alone 50-200 ng/ml, which were used in the studies
before 2000. Such high levels shut down leptin signaling pathways and explain the obesity-induced
infertility or subfertility. Indeed, a recent study by Wolodko et al details changes in ovarian gene
expression associated with leptin resistance caused by obesity(66). Collectively, these studies attest to
the importance of maintaining optimal levels of serum leptin for female reproductive competence.
Testicular Leptin Target cells: A little leptin goes a long way in the male. Zamorano et al (53)
first reported that Lepr mRNAs were expressed in the rat testes with subsequent studies identifying
t
all isoforms(204-206). In 1996, leptin treatment of ob/ob mice elevated serum FSH levels,
ip
increasing testicular and seminal vesicle weights and sperm counts(14).
cr
In the male, leptin is stimulatory before puberty. Suter et al(207) correlated the rise in serum
us
leptin levels with the pubertal nocturnal LH pulses in male monkeys showing that leptin is clearly
important in development. In isolated mouse seminiferous tubules, or 5-day old mouse Leydig cells,
an
leptin-induced an increase in phosphorylated STAT3 but not MAPK (208). In essence, in a number
of species, many studies of pubertal development in the male indicate that leptin is permissive and
M
stimulatory at the level of the hypothalamus and pituitary (23,64,106,149,153,154,207,209,210).

d
However, in the earlier section on sex differences in serum leptin levels, we reported that adult
e
males have relatively low serum leptin, when compared with females (46,109,110). Leptin levels in
pt
normal adult males are presumably limited by androgens, which inhibit secretion of adipocyte
leptin (46,109,110). This is fortuitous because, as will be shown in the following paragraphs, leptin
ce
inhibits testicular function if serum levels rise above a certain threshold.

Ac
With regard to Leydig cells, in vitro studies in cultured rat Leydig cells and a Leydig tumor line
demonstrated that obesity levels of leptin inhibited gonadotropin-mediated testosterone secretion
(23,210-213). The leptin suppression of testosterone was accompanied by parallel reductions in
androstenedione and a rise in progesterone and pregnenolone metabolites (23,212). In rat testicular
slices, leptin down-regulated testicular LEPRb (204,205,210,211). Researchers have pointed to the
positive correlation between high leptin in serum of obese men and reduced androgen levels (23).
After a report showed that circulating leptin could pass through the mouse blood-testes barrier
20
and thus enter the seminiferous tubules (214), researchers investigated leptin actions on
spermatogonia, spermatocytes and Sertoli cells. One report showed that leptin impaired mouse
sperm development and elements of the blood-testis barrier (215). Mice receiving 3 mg/kg leptin
for 2 weeks showed lower sperm counts (by 50.9%) and motility along with abnormal morphology.
Leptin also induced leakiness in the blood-testes barrier by reducing tight junction proteins
(occludin, claudin 5 and Zonula Occudins-1), actions that were prevented by inhibitors of leptin
signaling pathways: JAK2, PI3K, and MAPK. Collectively these studies explain the adverse
t
impact of high leptin on male mouse fertility and sperm counts. The actions on the tight junction
ip
proteins also pointed to leptin interactions with Sertoli cells, which form the tight junctions and the
cr
blood-testes barrier.
us
Sertoli cells are vital for the physical and nutritional support of spermatogenesis, producing
acetate from glucose. In isolated human Sertoli cells in culture (216), three concentrations of leptin
an
were tested, including adult male physiological levels (5 ng/mL) and levels found in obese (25
ng/mL) or morbidly obese (50 ng/mL) patients. Physiological concentration of leptin increased
M
glucose transporter 2 (GLUT2) proteins (216) indicating a route by which normal leptin levels may
maintain spermatogenesis. However, with obesity levels of leptin, there was a dose-dependent
d
decrease in acetate production, suggesting that rising leptin levels impair Sertoli cell nutritional
e
support for spermatocytes. These findings are important to our understanding of how too much
pt
leptin can interfere with Leydig cell production of testosterone and Sertoli cell's metabolic support
ce
of spermatogenesis.
Ac
Integrating Leptin Actions along the HPG Axis.
We have proposed an integrative hypothesis for female reproduction that supports a "leptin
permissive" partnership with the products of the HPG target cells to effect the optimal regulation of
reproduction (98)(Figure 2-4). We have further proposed that in the adult post-pubertal state, leptin's
support for reproduction is optimal only in a narrow concentration range, which has been shown to be
stimulatory to all target cells tested. Figures 2-4 summarize the major leptin target cells in the HPG
21
axis.
In females, the rise in leptin early in the cycle (from 14-20 ng/ml) is stimulated by estrogens from
the growing follicles (109) (Figure 4). Peak leptin levels synchronize with nocturnal LH pulses at
midcycle (111,112) because rising leptin has stimulated PMV neurons, which signal Kiss1 neurons
(117,121) (Figures 2 and 3) and subsequently, Kiss1 stimulates GnRH neurons (22,61). During the
follicular phase, rising estrogen from the developing follicles continues to stimulate the highest leptin
levels at the time of the LH surge (113), as it provides positive feedback to promote higher amplitude
t
and frequency GnRH pulses. Estrogen is also known to stimulate hormone and receptor expression in
ip
somatotropes, which promotes its role as a "somatogonadotrope", or secretion of GH as a co-
cr
gonadotropin (165,169-171,173) (Figure 3). And, leptin's actions at the level of the ovary may
us
enhance or permit the synergy between GH, IGF-1 and gonadotropins (187-190,194)(Figure 4).
In the pituitary, rising leptin levels stimulate LEPR expression (97) peaking at midcycle and
an
remaining high during the luteal phase. The combination of rising leptin and rapid GnRH pulses
stimulate GnRHR (98,99), thus facilitating the partnership between GnRH neurons and gonadotropes
M
by preparing them to receive the more rapid midcycle GnRH pulses. This molecular partnership
results in synchronized leptin-LH pulses and ovulation.

d
We have determined that leptin uses a post-transcriptional signaling pathway to increase pituitary
e
GnRHR levels and increase the sensitivity of gonadotropes to the more rapid GnRH pulses at
pt
midcycle(98,99). It is notable that Fshb mRNA also has MBEs in its 3'UTR, suggesting that cyclic
ce
de-repression of Fshb mRNA translation in response to leptin signaling could enhance translation
during the late luteal phase and may facilitate the early rise in FSH and the beginning of follicular
Ac
development for the next cycle.
22
In the ovary, we propose that rising serum leptin during the follicular phase can promote the
production of estrogen directly (202), or in the context of the FSH-growth factor synergy to produce
testosterone and estrogen. Hyperleptinemia caused by obesity may block estrogen production due to
leptin resistance(187-190,194). We also propose that the slightly higher luteal phase leptin levels (20
ng/ml) maintain corpus luteum function, possibly promoting the synergy between LH and growth
factor mediated estrogen and progesterone secretion. However, a higher leptin level in obesity will be
inhibitory(187-190,194). During the periovulatory period, we propose that normal leptin levels
t
promote meiotic progression of the oocytes after the LH surge. Thus, appropriate levels of leptin are
ip
required at all levels of the HPG as a permissive signal leading to successful reproduction.
cr
Data Availability
us
an
Some or all data generated or analyzed during this study are included in this published article or in
published articles that were reviewed by the authors. Also, because this is a review, data sharing of
M
findings by others may not be applicable for many sections as no datasets were generated or analyzed
during the writing of this review.

e d
pt
ce
Ac
23
References
1. Hileman SM, Pierroz DD, Flier JS. Leptin, nutrition, and reproduction: timing is
everything. J Clin Endocrinol Metab. 2000;85(2):804-807.
2. Kennedy GC, Mitra J. Body weight and food intake as initiating factors for puberty in
the rat. J Physiol. 1963;166:408-418.
3. Foster DL, Olster DH. Effect of restricted nutrition on puberty in the lamb: patterns of
t
tonic luteinizing hormone (LH) secretion and competency of the LH surge system.
ip
Endocrinology. 1985;116(1):375-381.
cr
4. Schillo KK. Effects of dietary energy on control of luteinizing hormone secretion in
us
cattle and sheep. J Anim Sci. 1992;70(4):1271-1282.
5. Wade GN, Schneider JE, Li HY. Control of fertility by metabolic cues. Am J Physiol.
an
1996;270(1 Pt 1):E1-19.
6. Gill CJ, Rissman EF. Female sexual behavior is inhibited by short- and long-term
M
food restriction. Physiol Behav. 1997;61(3):387-394.

d
7. Hamilton GD, Bronson FH. Food restriction and reproductive development: male and
e
female mice and male rats. Am J Physiol. 1986;250(3 Pt 2):R370-376.

pt
8. Cameron JL, Nosbisch C. Suppression of pulsatile luteinizing hormone and

ce
testosterone secretion during short term food restriction in the adult male rhesus
monkey (Macaca mulatta). Endocrinology. 1991;128(3):1532-1540.

Ac
9. Cameron JL, Weltzin TE, McConaha C, Helmreich DL, Kaye WH. Slowing of
pulsatile luteinizing hormone secretion in men after forty-eight hours of fasting. J
Clin Endocrinol Metab. 1991;73(1):35-41.
10. Foster DL, Nagatani S. Physiological perspectives on leptin as a regulator of
reproduction: role in timing puberty. Biol Reprod. 1999;60(2):205-215.
24
11. Considine RV, Sinha MK, Heiman ML, Kriauciunas A, Stephens TW, Nyce MR,
Ohannesian JP, Marco CC, McKee LJ, Bauer TL, et al. Serum immunoreactive-leptin
concentrations in normal-weight and obese humans. The New England journal of
medicine. 1996;334(5):292-295.
12. Ruhl CE, Harris TB, Ding J, Goodpaster BH, Kanaya AM, Kritchevsky SB,
Simonsick EM, Tylavsky FA, Everhart JE. Body mass index and serum leptin
concentration independently estimate percentage body fat in older adults. Am J Clin
t
ip
Nutr. 2007;85(4):1121-1126.
cr
13. Zhang Y, Proenca R, Maffei M, Barone M, Leopold L, Friedman JM. Positional
cloning of the mouse obese gene and its human homologue. Nature.
1994;372(6505):425-432.
us
an
14. Barash IA, Cheung CC, Weigle DS, Ren H, Kabigting EB, Kuijper JL, Clifton DK,
Steiner RA. Leptin is a metabolic signal to the reproductive system. Endocrinology.

M
1996;137(7):3144-3147.
15. Chehab FF, Lim ME, Lu R. Correction of the sterility defect in homozygous obese
e d
female mice by treatment with the human recombinant leptin. Nat Genet.
pt
1996;12(3):318-320.
16. Ahima RS, Dushay J, Flier SN, Prabakaran D, Flier JS. Leptin accelerates the onset of
ce
puberty in normal female mice. J Clin Invest. 1997;99(3):391-395.

Ac
17. Kiess W, Blum WF, Aubert ML. Leptin, puberty and reproductive function: lessons
from animal studies and observations in humans. European Journal of Endocrinology
/ European Federation of Endocrine Societies. 1998;138(1):26-29.
18. Casanueva FF, Dieguez C. Neuroendocrine regulation and actions of leptin. Frontiers
in Neuroendocrinology. 1999;20(4):317-363.
19. Ahima RS, Flier JS. Leptin. Annu Rev Physiol. 2000;62:413-437.
25
20. Chou SH, Mantzoros C. 20 years of leptin: role of leptin in human reproductive
disorders. J Endocrinol. 2014;223(1):T49-62.
21. Elias CF. Leptin action in pubertal development: recent advances and unanswered
questions. Trends in Endocrinology and Metabolism: TEM. 2012;23(1):9-15.
22. Elias CF, Purohit D. Leptin signaling and circuits in puberty and fertility. Cell Mol
Life Sci. 2013;70(5):841-862.
23. Caprio M, Fabbrini E, Isidori AM, Aversa A, Fabbri A. Leptin in reproduction.
t
ip
Trends Endocrinol Metab. 2001;12(2):65-72.
cr
24. Pralong FP, Gaillard RC. Neuroendocrine effects of leptin. Pituitary. 2001;4(1-2):25-
32.
25.
us
Bluher S, Mantzoros CS. Leptin in reproduction. Current opinion in endocrinology,
an
diabetes, and obesity. 2007;14(6):458-464.
26. Mantzoros CS, Magkos F, Brinkoetter M, Sienkiewicz E, Dardeno TA, Kim SY,
M
Hamnvik OP, Koniaris A. Leptin in human physiology and pathophysiology.
American Journal of Physiology Endocrinology and metabolism. 2011;301(4):E567-

e d
584.
pt
27. Paz-Filho G, Wong ML, Licinio J. Ten years of leptin replacement therapy. Obes Rev.
2011;12(5):e315-323. doi: 310.1111/j.1467-1789X.2010.00840.x. Epub 02011 Mar

ce
00817.
Ac
28. Coleman DL. A historical perspective on leptin. Nat Med. 2010;16(10):1097-1099.
29. Ingalls AM, Dickie MM, Snell GD. Obese, a new mutation in the house mouse. J
Hered. 1950;41(12):317-318.
30. Hummel KP, Dickie MM, Coleman DL. Diabetes, a new mutation in the mouse.
Science. 1966;153(3740):1127-1128.
26
31. Swerdloff RS, Batt RA, Bray GA. Reproductive hormonal function in the genetically
obese (ob/ob) mouse. Endocrinology. 1976;98(6):1359-1364.
32. Coleman DL, Hummel KP. Effects of parabiosis of normal with genetically diabetic
mice. Am J Physiol. 1969;217(5):1298-1304.
33. Coleman DL. Effects of parabiosis of obese with diabetes and normal mice.
Diabetologia. 1973;9(4):294-298.
34. Lee GH, Proenca R, Montez JM, Carroll KM, Darvishzadeh JG, Lee JI, Friedman JM.
t
ip
Abnormal splicing of the leptin receptor in diabetic mice. Nature.
cr
1996;379(6566):632-635.
35. Chen H, Charlat O, Tartaglia LA, Woolf EA, Weng X, Ellis SJ, Lakey ND, Culpepper
us
J, Moore KJ, Breitbart RE, Duyk GM, Tepper RI, Morgenstern JP. Evidence that the
an
diabetes gene encodes the leptin receptor: identification of a mutation in the leptin
receptor gene in db/db mice. Cell. 1996;84(3):491-495.

M
36. Ghilardi N, Ziegler S, Wiestner A, Stoffel R, Heim MH, Skoda RC. Defective STAT
signaling by the leptin receptor in diabetic mice. Proc Natl Acad Sci U S A.
e d
1996;93(13):6231-6235.
pt
37. Tartaglia LA, Dembski M, Weng X, Deng N, Culpepper J, Devos R, Richards GJ,
Campfield LA, Clark FT, Deeds J, Muir C, Sanker S, Moriarty A, Moore KJ, Smutko
ce
JS, Mays GG, Wool EA, Monroe CA, Tepper RI. Identification and expression
Ac
cloning of a leptin receptor, OB-R. Cell. 1995;83(7):1263-1271.
38. Cammisotto PG, Gelinas Y, Deshaies Y, Bukowiecki LJ. Regulation of leptin
secretion from white adipocytes by insulin, glycolytic substrates, and amino acids. Am
J Physiol Endocrinol Metab. 2005;289(1):E166-171.
27
39. Saladin R, De Vos P, Guerre-Millo M, Leturque A, Girard J, Staels B, Auwerx J.
Transient increase in obese gene expression after food intake or insulin
administration. Nature. 1995;377(6549):527-529.
40. Menendez C, Baldelli R, Camina JP, Escudero B, Peino R, Dieguez C, Casanueva FF.
TSH stimulates leptin secretion by a direct effect on adipocytes. The Journal of
endocrinology. 2003;176(1):7-12.
41. Isozaki O, Tsushima T, Miyakawa M, Demura H, Seki H. Interaction between leptin
t
ip
and growth hormone (GH)/IGF-I axis. Endocr J. 1999;46 Suppl:S17-24.
cr
42. Isozaki O, Tsushima T, Miyakawa M, Nozoe Y, Demura H, Seki H. Growth hormone
directly inhibits leptin gene expression in visceral fat tissue in fatty Zucker rats. J
Endocrinol. 1999;161(3):511-516.
us
an
43. Fungfuang W, Nakada T, Nakao N, Terada M, Yokosuka M, Gizurarson S, Hau J,
Moon C, Saito TR. Serum leptin concentrations, leptin mRNA expression, and food
M
intake during the estrous cycle in rats. Lab Anim Res. 2013;29(1):1-6.
44. Fungfuang W, Terada M, Komatsu N, Moon C, Saito TR. Effects of estrogen on food
e d
intake, serum leptin levels and leptin mRNA expression in adipose tissue of female
pt
rats. Lab Anim Res. 2013;29(3):168-173.
45. Jenks MZ, Fairfield HE, Johnson EC, Morrison RF, Muday GK. Sex Steroid
ce
Hormones Regulate Leptin Transcript Accumulation and Protein Secretion in 3T3-L1

Ac
Cells. Sci Rep. 2017;7(1):8232.
46. Luukkaa V, Pesonen U, Huhtaniemi I, Lehtonen A, Tilvis R, Tuomilehto J, Koulu M,
Huupponen R. Inverse correlation between serum testosterone and leptin in men. J
Clin Endocrinol Metab. 1998;83(9):3243-3246.
28
47. Peelman F, Zabeau L, Moharana K, Savvides SN, Tavernier J. 20 years of leptin:
insights into signaling assemblies of the leptin receptor. J Endocrinol.
2014;223(1):T9-23.
48. Wauman J, Zabeau L, Tavernier J. The Leptin Receptor Complex: Heavier Than
Expected? Front Endocrinol (Lausanne). 2017;8:30.
49. Baumann H, Morella KK, White DW, Dembski M, Bailon PS, Kim H, Lai CF,
Tartaglia LA. The full-length leptin receptor has signaling capabilities of interleukin
t
ip
6-type cytokine receptors. Proc Natl Acad Sci U S A. 1996;93(16):8374-8378.
cr
50. Chua SC, Jr., Koutras IK, Han L, Liu SM, Kay J, Young SJ, Chung WK, Leibel RL.
Fine structure of the murine leptin receptor gene: splice site suppression is required to
us
form two alternatively spliced transcripts. Genomics. 1997;45(2):264-270.
an
51. White DW, Kuropatwinski KK, Devos R, Baumann H, Tartaglia LA. Leptin receptor
(OB-R) signaling. Cytoplasmic domain mutational analysis and evidence for receptor
M
homo-oligomerization. The Journal of biological chemistry. 1997;272(7):4065-4071.
52. Yamashita T, Murakami T, Otani S, Kuwajima M, Shima K. Leptin receptor signal

e d
transduction: OBRa and OBRb of fa type. Biochemical and biophysical research

pt
communications. 1998;246(3):752-759.
53. Zamorano PL, Mahesh VB, De Sevilla LM, Chorich LP, Bhat GK, Brann DW.
ce
Expression and localization of the leptin receptor in endocrine and neuroendocrine

Ac
tissues of the rat. Neuroendocrinology. 1997;65(3):223-228.
54. Patterson CM, Villanueva EC, Greenwald-Yarnell M, Rajala M, Gonzalez IE, Saini
N, Jones J, Myers MG, Jr. Leptin action via LepR-b Tyr1077 contributes to the
control of energy balance and female reproduction. Mol Metab. 2012;1(1-2):61-69.
55. Gao Q, Wolfgang MJ, Neschen S, Morino K, Horvath TL, Shulman GI, Fu XY.
Disruption of neural signal transducer and activator of transcription 3 causes obesity,
29
diabetes, infertility, and thermal dysregulation. Proc Natl Acad Sci U S A.
2004;101(13):4661-4666.
56. Singireddy AV, Inglis MA, Zuure WA, Kim JS, Anderson GM. Neither signal
transducer and activator of transcription 3 (STAT3) or STAT5 signaling pathways are
required for leptin's effects on fertility in mice. Endocrinology. 2013;154(7):2434-
2445.
57. Fruhbeck G. Intracellular signalling pathways activated by leptin. Biochem J.
t
ip
2006;393(Pt 1):7-20.
cr
58. Bjorbaek C, Buchholz RM, Davis SM, Bates SH, Pierroz DD, Gu H, Neel BG, Myers
MG, Jr., Flier JS. Divergent roles of SHP-2 in ERK activation by leptin receptors. J
Biol Chem. 2001;276(7):4747-4755.

us
an
59. Myers MG, Jr. Leptin receptor signaling and the regulation of mammalian
physiology. Recent Prog Horm Res. 2004;59:287-304.

M
60. Niswender KD, Morton GJ, Stearns WH, Rhodes CJ, Myers MG, Jr., Schwartz MW.
Intracellular signalling. Key enzyme in leptin-induced anorexia. Nature.

e d
2001;413(6858):794-795.
pt
61. Elias CF. Leptin action in pubertal development: recent advances and unanswered
questions. Trends Endocrinol Metab. 2012;23(1):9-15.

ce
62. Roa J, Garcia-Galiano D, Castellano JM, Gaytan F, Pinilla L, Tena-Sempere M.

Ac
Metabolic control of puberty onset: new players, new mechanisms. Mol Cell
Endocrinol. 2010;324(1-2):87-94.
63. Roa J, Garcia-Galiano D, Varela L, Sanchez-Garrido MA, Pineda R, Castellano JM,
Ruiz-Pino F, Romero M, Aguilar E, Lopez M, Gaytan F, Dieguez C, Pinilla L, Tena-
Sempere M. The mammalian target of rapamycin as novel central regulator of puberty
30
onset via modulation of hypothalamic Kiss1 system. Endocrinology.
2009;150(11):5016-5026.
64. Yu WH, Walczewska A, Karanth S, McCann SM. Nitric oxide mediates leptin-
induced luteinizing hormone-releasing hormone (LHRH) and LHRH and leptin-
induced LH release from the pituitary gland. Endocrinology. 1997;138(11):5055-
5058.
65. Riad-Gabriel MG, Jinagouda SD, Sharma A, Boyadjian R, Saad MF. Changes in
t
ip
plasma leptin during the menstrual cycle. Eur J Endocrinol. 1998;139(5):528-531.
cr
66. Wolodko K, Walewska E, Adamowski M, Castillo-Fernandez J, Kelsey G, Galvao A.
Leptin Resistance in the Ovary of Obese Mice is Associated with Profound Changes
us
in the Transcriptome of Cumulus Cells. Cell Physiol Biochem. 2020;54(3):417-437.
an
67. Bjorbaek C, Uotani S, da Silva B, Flier JS. Divergent signaling capacities of the long
and short isoforms of the leptin receptor. J Biol Chem. 1997;272(51):32686-32695.

M
68. Odle AK, Haney A, Allensworth-James M, Akhter N, Childs GV. Adipocyte vs
pituitary leptin in the regulation of pituitary hormones: Somatotropes develop

e d
normally in the absence of circulating leptin. Endocrinology. 2014;155(Nov):4316-

pt
4328.
69. Chehab FF, Mounzih K, Lu R, Lim ME. Early onset of reproductive function in
ce
normal female mice treated with leptin. Science. 1997;275(5296):88-90.

Ac
70. Cheung CC, Thornton JE, Kuijper JL, Weigle DS, Clifton DK, Steiner RA. Leptin is a
metabolic gate for the onset of puberty in the female rat. Endocrinology.
1997;138(2):855-858.
71. Bronson FH. Food-restricted, prepubertal, female rats: rapid recovery of luteinizing
hormone pulsing with excess food, and full recovery of pubertal development with
gonadotropin-releasing hormone. Endocrinology. 1986;118(6):2483-2487.
31
72. Bronson FH. Puberty in female mice is not associated with increases in either body fat
or leptin. Endocrinology. 2001;142(11):4758-4761.
73. Cheung CC, Thornton JE, Nurani SD, Clifton DK, Steiner RA. A reassessment of
leptin's role in triggering the onset of puberty in the rat and mouse.
Neuroendocrinology. 2001;74(1):12-21.
74. Mann DR, Akinbami MA, Gould KG, Castracane VD. Leptin and thyroxine during
sexual development in male monkeys: effect of neonatal gonadotropin-releasing
t
ip
hormone antagonist treatment and delayed puberty on the developmental pattern of
cr
leptin and thyroxine secretion. Eur J Endocrinol. 2002;146(6):891-898.
75. Mann DR, Bhat GK, Ramaswamy S, Stah CD, Plant TM. Regulation of circulating
us
leptin and its soluble receptor during pubertal development in the male rhesus monkey
an
(Macaca mulatta). Endocrine. 2007;31(2):125-129.
76. Plant TM, Durrant AR. Circulating leptin does not appear to provide a signal for
M
triggering the initiation of puberty in the male rhesus monkey (Macaca mulatta).
Endocrinology. 1997;138(10):4505-4508.
e d
77. Urbanski HF, Pau KY. A biphasic developmental pattern of circulating leptin in the
pt
male rhesus macaque (Macaca mulatta). Endocrinology. 1998;139(5):2284-2286.
78. Jaquet D, Leger J, Levy-Marchal C, Oury JF, Czernichow P. Ontogeny of leptin in

ce
human fetuses and newborns: effect of intrauterine growth retardation on serum leptin
Ac
concentrations. J Clin Endocrinol Metab. 1998;83(4):1243-1246.
79. Cetin I, Morpurgo PS, Radaelli T, Taricco E, Cortelazzi D, Bellotti M, Pardi G, Beck-
Peccoz P. Fetal plasma leptin concentrations: relationship with different intrauterine
growth patterns from 19 weeks to term. Pediatr Res. 2000;48(5):646-651.
32
80. Ahima RS, Prabakaran D, Flier JS. Postnatal leptin surge and regulation of circadian
rhythm of leptin by feeding. Implications for energy homeostasis and neuroendocrine
function. J Clin Invest. 1998;101(5):1020-1027.
81. Devaskar SU, Ollesch C, Rajakumar RA, Rajakumar PA. Developmental changes in
ob gene expression and circulating leptin peptide concentrations. Biochem Biophys
Res Commun. 1997;238(1):44-47.
82. Delahaye F, Breton C, Risold PY, Enache M, Dutriez-Casteloot I, Laborie C, Lesage
t
ip
J, Vieau D. Maternal perinatal undernutrition drastically reduces postnatal leptin surge
cr
and affects the development of arcuate nucleus proopiomelanocortin neurons in
neonatal male rat pups. Endocrinology. 2008;149(2):470-475.
83.
us
Vickers MH, Gluckman PD, Coveny AH, Hofman PL, Cutfield WS, Gertler A, Breier
an
BH, Harris M. Neonatal leptin treatment reverses developmental programming.
Endocrinology. 2005;146(10):4211-4216.
M
84. Yura S, Itoh H, Sagawa N, Yamamoto H, Masuzaki H, Nakao K, Kawamura M,
Takemura M, Kakui K, Ogawa Y, Fujii S. Role of premature leptin surge in obesity

e d
resulting from intrauterine undernutrition. Cell Metab. 2005;1(6):371-378.

pt
85. Mistry AM, Swick A, Romsos DR. Leptin alters metabolic rates before acquisition of
its anorectic effect in developing neonatal mice. Am J Physiol. 1999;277(3):R742-

ce
747.
Ac
86. Proulx K, Richard D, Walker CD. Leptin regulates appetite-related neuropeptides in
the hypothalamus of developing rats without affecting food intake. Endocrinology.
2002;143(12):4683-4692.
87. Bouret SG, Draper SJ, Simerly RB. Trophic action of leptin on hypothalamic neurons
that regulate feeding. Science. 2004;304(5667):108-110.
33
88. Bouret SG, Simerly RB. Minireview: Leptin and development of hypothalamic
feeding circuits. Endocrinology. 2004;145(6):2621-2626.
89. Ramos-Lobo AM, Teixeira PD, Furigo IC, Melo HM, de MLESN, De Felice FG,
Donato J, Jr. Long-term consequences of the absence of leptin signaling in early life.
Elife. 2019;8.
90. Attig L, Larcher T, Gertler A, Abdennebi-Najar L, Djiane J. Postnatal leptin is
necessary for maturation of numerous organs in newborn rats. Organogenesis.
t
ip
2011;7(2):88-94.
cr
91. Coupe B, Amarger V, Grit I, Benani A, Parnet P. Nutritional programming affects
hypothalamic organization and early response to leptin. Endocrinology.
2010;151(2):702-713.
us
an
92. Lopez-Gallardo M, Anton-Fernandez A, Llorente R, Mela V, Llorente-Berzal A,
Prada C, Viveros MP. Neonatal Treatment with a Pegylated Leptin Antagonist

M
Induces Sexually Dimorphic Effects on Neurones and Glial Cells, and on Markers of
Synaptic Plasticity in the Developing Rat Hippocampal Formation. J

e d
Neuroendocrinol. 2015;27(8):658-669.
pt
93. Leonhardt M, Lesage J, Croix D, Dutriez-Casteloot I, Beauvillain JC, Dupouy JP.
Effects of perinatal maternal food restriction on pituitary-gonadal axis and plasma

ce
leptin level in rat pup at birth and weaning and on timing of puberty. Biol Reprod.
Ac
2003;68(2):390-400.
94. Mela V, Diaz F, Lopez-Rodriguez AB, Vazquez MJ, Gertler A, Argente J, Tena-
Sempere M, Viveros MP, Chowen JA. Blockage of the Neonatal Leptin Surge Affects
the Gene Expression of Growth Factors, Glial Proteins, and Neuropeptides Involved
in the Control of Metabolism and Reproduction in Peripubertal Male and Female
Rats. Endocrinology. 2015;156(7):2571-2581.
34
95. Castellano JM, Bentsen AH, Sanchez-Garrido MA, Ruiz-Pino F, Romero M, Garcia-
Galiano D, Aguilar E, Pinilla L, Dieguez C, Mikkelsen JD, Tena-Sempere M. Early
metabolic programming of puberty onset: impact of changes in postnatal feeding and
rearing conditions on the timing of puberty and development of the hypothalamic
kisspeptin system. Endocrinology. 2011;152(9):3396-3408.
96. Iwasa T, Matsuzaki T, Murakami M, Kinouchi R, Gereltsetseg G, Fujisawa S,
Kuwahara A, Yasui T, Irahara M. Sensitivities of mRNA expression levels of Kiss1
t
ip
and its receptor, Kiss1r, to nutritional status are changed during the developmental
cr
period in female rats. J Endocrinol. 2010;207(2):195-202.
97. Akhter N, CarlLee T, Syed MM, Odle AK, Cozart MA, Haney AC, Allensworth-
us
James ML, Benes H, Childs GV. Selective deletion of leptin receptors in
an
gonadotropes reveals activin and GnRH-binding sites as leptin targets in support of
fertility. Endocrinology. 2014;155(10):4027-4042.

M
98. Odle A, Akhter N, Syed M, Allensworth-James M, Beneš H, Melgar-Castillo A,
MacNicol M, MacNicol A, Childs GV. Hypothesis and Theory: Leptin Regulation of

e d
Gonadotrope Gonadotropin Releasing Hormone Receptors (GnRHR) as a Metabolic

pt
Checkpoint and Gateway to Reproductive Competence. Front Endocrinol (Lausanne).
2017.
ce
99. Odle AK, Benes H, Melgar Castillo A, Akhter N, Syed M, Haney A, Allensworth-
Ac
James M, Hardy L, Winter B, Manoharan R, Syed R, MacNicol MC, MacNicol AM,
Childs GV. Association of Gnrhr mRNA With the Stem Cell Determinant Musashi: A
Mechanism for Leptin-Mediated Modulation of GnRHR Expression. Endocrinology.
2018;159(2):883-894.
35
100. Wen S, Ai W, Alim Z, Boehm U. Embryonic gonadotropin-releasing hormone
signaling is necessary for maturation of the male reproductive axis. Proc Natl Acad
Sci U S A. 2010;107(37):16372-16377.
101. Dohler KD, Wuttke W. Changes with age in levels of serum gonadotropins, prolactin
and gonadal steroids in prepubertal male and female rats. Endocrinology.
1975;97(4):898-907.
102. Bjelobaba I, Janjic MM, Kucka M, Stojilkovic SS. Cell Type-Specific Sexual
t
ip
Dimorphism in Rat Pituitary Gene Expression During Maturation. Biol Reprod.
cr
2015;93(1):21.
103. Wilson ME, Handa RJ. Ontogeny of gene expression in the gonadotroph of the
us
developing female rat. Biol Reprod. 1997;56(2):563-568.
an
104. Wilson ME, Handa RJ. Activin subunit, follistatin, and activin receptor gene
expression in the prepubertal female rat pituitary. Biol Reprod. 1998;59(2):278-283.

M
105. Suter KJ, Pohl CR, Plant TM. The pattern and tempo of the pubertal reaugmentation
of open-loop pulsatile gonadotropin-releasing hormone release assessed indirectly in

e d
the male rhesus monkey (Macaca mulatta). Endocrinology. 1998;139(6):2774-2783.

pt
106. Nagatani S, Guthikonda P, Foster DL. Appearance of a nocturnal peak of leptin
secretion in the pubertal rat. Horm Behav. 2000;37(4):345-352.

ce
107. Garcia-Mayor RV, Andrade MA, Rios M, Lage M, Dieguez C, Casanueva FF. Serum
Ac
leptin levels in normal children: relationship to age, gender, body mass index,
pituitary-gonadal hormones, and pubertal stage. J Clin Endocrinol Metab.
1997;82(9):2849-2855.
108. Isidori AM, Strollo F, More M, Caprio M, Aversa A, Moretti C, Frajese G, Riondino
G, Fabbri A. Leptin and aging: correlation with endocrine changes in male and female
36
healthy adult populations of different body weights. J Clin Endocrinol Metab.
2000;85(5):1954-1962.
109. Wauters M, Considine RV, Van Gaal LF. Human leptin: from an adipocyte hormone
to an endocrine mediator. Eur J Endocrinol. 2000;143(3):293-311.
110. Wabitsch M, Blum WF, Muche R, Braun M, Hube F, Rascher W, Heinze E, Teller W,
Hauner H. Contribution of androgens to the gender difference in leptin production in
obese children and adolescents. J Clin Invest. 1997;100(4):808-813.
t
ip
111. Licinio J, Negrao AB, Mantzoros C, Kaklamani V, Wong ML, Bongiorno PB, Mulla
cr
A, Cearnal L, Veldhuis JD, Flier JS, McCann SM, Gold PW. Synchronicity of
frequently sampled, 24-h concentrations of circulating leptin, luteinizing hormone,
us
and estradiol in healthy women. Proc Natl Acad Sci U S A. 1998;95(5):2541-2546.
an
112. Sir-Petermann T, Piwonka V, Perez F, Maliqueo M, Recabarren SE, Wildt L. Are
circulating leptin and luteinizing hormone synchronized in patients with polycystic

M
ovary syndrome? Hum Reprod. 1999;14(6):1435-1439.
113. Ahrens K, Mumford SL, Schliep KC, Kissell KA, Perkins NJ, Wactawski-Wende J,
e d
Schisterman EF. Serum leptin levels and reproductive function during the menstrual
pt
cycle. Am J Obstet Gynecol. 2014;210(3):248 e241-249.
114. McMinn JE, Liu SM, Liu H, Dragatsis I, Dietrich P, Ludwig T, Boozer CN, Chua SC,
ce
Jr. Neuronal deletion of Lepr elicits diabesity in mice without affecting cold tolerance
Ac
or fertility. Am J Physiol Endocrinol Metab. 2005;289(3):E403-411.
115. Chan JL, Mantzoros CS. Leptin and the hypothalamic-pituitary regulation of the
gonadotropin-gonadal axis. Pituitary. 2001;4(1-2):87-92.
116. Quennell JH, Mulligan AC, Tups A, Liu X, Phipps SJ, Kemp CJ, Herbison AE,
Grattan DR, Anderson GM. Leptin indirectly regulates gonadotropin-releasing
hormone neuronal function. Endocrinology. 2009;150(6):2805-2812.
37
117. Donato J, Jr., Cravo RM, Frazao R, Gautron L, Scott MM, Lachey J, Castro IA,
Margatho LO, Lee S, Lee C, Richardson JA, Friedman J, Chua S, Jr., Coppari R,
Zigman JM, Elmquist JK, Elias CF. Leptin's effect on puberty in mice is relayed by
the ventral premammillary nucleus and does not require signaling in Kiss1 neurons. J
Clin Invest. 2011;121(1):355-368.
118. Navarro VM. Metabolic regulation of kisspeptin - the link between energy balance
and reproduction. Nat Rev Endocrinol. 2020;16(8):407-420.
t
ip
119. Evans MC, Anderson GM. Integration of Circadian and Metabolic Control of
cr
Reproductive Function. Endocrinology. 2018;159(11):3661-3673.
120. Donato J, Jr., Elias CF. The ventral premammillary nucleus links metabolic cues and
us
reproduction. Front Endocrinol (Lausanne). 2011;2:57.
an
121. Donato J, Jr., Silva RJ, Sita LV, Lee S, Lee C, Lacchini S, Bittencourt JC, Franci CR,
Canteras NS, Elias CF. The ventral premammillary nucleus links fasting-induced
M
changes in leptin levels and coordinated luteinizing hormone secretion. The Journal of
neuroscience : the official journal of the Society for Neuroscience. 2009;29(16):5240-

e d
5250.
pt
122. Mahany EB, Han X, Borges BC, da Silveira Cruz-Machado S, Allen SJ, Garcia-
Galiano D, Hoenerhoff MJ, Bellefontaine NH, Elias CF. Obesity and High-Fat Diet
ce
Induce Distinct Changes in Placental Gene Expression and Pregnancy Outcome.

Ac
Endocrinology. 2018;159(4):1718-1733.
123. Ross RA, Leon S, Madara JC, Schafer D, Fergani C, Maguire CA, Verstegen AM,
Brengle E, Kong D, Herbison AE, Kaiser UB, Lowell BB, Navarro VM. PACAP
neurons in the ventral premammillary nucleus regulate reproductive function in the
female mouse. Elife. 2018;7.
38
124. Petrine JCP, Franci CR, Del Bianco-Borges B. Leptin actions through the nitrergic
system to modulate the hypothalamic expression of the kiss1 mRNA in the female rat.
Brain Res. 2020;1728:146574.
125. Zuure WA, Roberts AL, Quennell JH, Anderson GM. Leptin signaling in GABA
neurons, but not glutamate neurons, is required for reproductive function. J Neurosci.
2013;33(45):17874-17883.
126. Martin C, Navarro VM, Simavli S, Vong L, Carroll RS, Lowell BB, Kaiser UB.
t
ip
Leptin-responsive GABAergic neurons regulate fertility through pathways that result
cr
in reduced kisspeptinergic tone. J Neurosci. 2014;34(17):6047-6056.
127. Padilla SL, Qiu J, Nestor CC, Zhang C, Smith AW, Whiddon BB, Ronnekleiv OK,
us
Kelly MJ, Palmiter RD. AgRP to Kiss1 neuron signaling links nutritional state and
an
fertility. Proc Natl Acad Sci U S A. 2017;114(9):2413-2418.
128. Wu Q, Whiddon BB, Palmiter RD. Ablation of neurons expressing agouti-related

M
protein, but not melanin concentrating hormone, in leptin-deficient mice restores
metabolic functions and fertility. Proc Natl Acad Sci U S A. 2012;109(8):3155-3160.

e d
129. Egan OK, Inglis MA, Anderson GM. Leptin Signaling in AgRP Neurons Modulates
pt
Puberty Onset and Adult Fertility in Mice. J Neurosci. 2017;37(14):3875-3886.
130. Jin L, Burguera BG, Couce ME, Scheithauer BW, Lamsan J, Eberhardt NL, Kulig E,
ce
Lloyd RV. Leptin and leptin receptor expression in normal and neoplastic human
Ac
pituitary: evidence of a regulatory role for leptin on pituitary cell proliferation. J Clin
Endocrinol Metab. 1999;84(8):2903-2911.
131. Jin L, Zhang S, Burguera BG, Couce ME, Osamura RY, Kulig E, Lloyd RV. Leptin
and leptin receptor expression in rat and mouse pituitary cells. Endocrinology.
2000;141(1):333-339.
39
132. Iqbal J, Kurose Y, Canny B, Clarke IJ. Effects of central infusion of ghrelin on food
intake and plasma levels of growth hormone, luteinizing hormone, prolactin, and
cortisol secretion in sheep. Endocrinology. 2006;147(1):510-519.
133. Iqbal J, Pompolo S, Considine RV, Clarke IJ. Localization of leptin receptor-like
immunoreactivity in the corticotropes, somatotropes, and gonadotropes in the ovine
anterior pituitary. Endocrinology. 2000;141(4):1515-1520.
134. Lloyd RV, Jin L, Qian X, Zhang S, Scheithauer BW. Nitric oxide synthase in the
t
ip
human pituitary gland. Am J Pathol. 1995;146(1):86-94.
cr
135. Lloyd RV, Jin L, Tsumanuma I, Vidal S, Kovacs K, Horvath E, Scheithauer BW,
Couce ME, Burguera B. Leptin and leptin receptor in anterior pituitary function.
Pituitary. 2001;4(1-2):33-47.
us
an
136. Giusti M, Bocca L, Florio T, Corsaro A, Spaziante R, Schettini G, Minuto F. In vitro
effect of human recombinant leptin and expression of leptin receptors on growth

M
hormone-secreting human pituitary adenomas. Clin Endocrinol (Oxf).
2002;57(4):449-455.
e d
137. Sone M, Osamura RY. Leptin and the pituitary. Pituitary. 2001;4(1-2):15-23.
pt
138. Mann DR, Plant TM. Leptin and pubertal development. Semin Reprod Med.
2002;20(2):93-102.
ce
139. Urbanski HF. Leptin and puberty. Trends Endocrinol Metab. 2001;12(10):428-429.
Ac
140. Yura S, Ogawa Y, Sagawa N, Masuzaki H, Itoh H, Ebihara K, Aizawa-Abe M, Fujii
S, Nakao K. Accelerated puberty and late-onset hypothalamic hypogonadism in
female transgenic skinny mice overexpressing leptin. J Clin Invest. 2000;105(6):749-
755.
141. Kaminski T, Smolinska N, Gajewska A, Siawrys G, Okrasa S, Kochman K, Przala J.
Leptin and long form of leptin receptor genes expression in the hypothalamus and
40
pituitary during the luteal phase and early pregnancy in pigs. J Physiol Pharmacol.
2006;57(1):95-108.
142. Carro E, Pinilla L, Seoane LM, Considine RV, Aguilar E, Casanueva FF, Dieguez C.
Influence of endogenous leptin tone on the estrous cycle and luteinizing hormone
pulsatility in female rats. Neuroendocrinology. 1997;66(6):375-377.
143. De Biasi SN, Apfelbaum LI, Apfelbaum ME. In vitro effect of leptin on LH release
by anterior pituitary glands from female rats at the time of spontaneous and steroid-
t
ip
induced LH surge. Eur J Endocrinol. 2001;145(5):659-665.
cr
144. Schneider JE, Goldman MD, Tang S, Bean B, Ji H, Friedman MI. Leptin indirectly
affects estrous cycles by increasing metabolic fuel oxidation. Hormones and behavior.
1998;33(3):217-228.
us
an
145. Ogura K, Irahara M, Kiyokawa M, Tezuka M, Matsuzaki T, Yasui T, Kamada M,
Aono T. Effects of leptin on secretion of LH and FSH from primary cultured female
M
rat pituitary cells. Eur J Endocrinol. 2001;144(6):653-658.
146. Spicer LJ. Leptin: a possible metabolic signal affecting reproduction. Domest Anim
e d
Endocrinol. 2001;21(4):251-270.
pt
147. Tezuka M, Irahara M, Ogura K, Kiyokawa M, Tamura T, Matsuzaki T, Yasui T,
Aono T. Effects of leptin on gonadotropin secretion in juvenile female rat pituitary

ce
cells. Eur J Endocrinol. 2002;146(2):261-266.

Ac
148. Swerdloff RS, Peterson M, Vera A, Batt RA, Heber D, Bray GA. The hypothalamic-
pituitary axis in genetically obese (ob/ob) mice: response to luteinizing hormone-
releasing hormone. Endocrinology. 1978;103(2):542-547.
149. Yu WH, Kimura M, Walczewska A, Karanth S, McCann SM. Role of leptin in
hypothalamic-pituitary function. Proc Natl Acad Sci U S A. 1997;94(3):1023-1028.
41
150. Ahima RS, Prabakaran D, Mantzoros C, Qu D, Lowell B, Maratos-Flier E, Flier JS.
Role of leptin in the neuroendocrine response to fasting. Nature. 1996;382(6588):250-
252.
151. Finn PD, Cunningham MJ, Pau KY, Spies HG, Clifton DK, Steiner RA. The
stimulatory effect of leptin on the neuroendocrine reproductive axis of the monkey.
Endocrinology. 1998;139(11):4652-4662.
152. Gonzalez LC, Pinilla L, Tena-Sempere M, Aguilar E. Leptin(116-130) stimulates
t
ip
prolactin and luteinizing hormone secretion in fasted adult male rats.
cr
153. Nagatani S, Guthikonda P, Thompson RC, Tsukamura H, Maeda KI, Foster DL.
us
Evidence for GnRH regulation by leptin: leptin administration prevents reduced
an
pulsatile LH secretion during fasting. Neuroendocrinology. 1998;67(6):370-376.
154. Nagatani S, Zeng Y, Keisler DH, Foster DL, Jaffe CA. Leptin regulates pulsatile
M
luteinizing hormone and growth hormone secretion in the sheep. Endocrinology.
2000;141(11):3965-3975.
e d
155. Schneider JE, Blum RM, Wade GN. Metabolic control of food intake and estrous
pt
cycles in syrian hamsters. I. Plasma insulin and leptin. Am J Physiol Regul Integr
Comp Physiol. 2000;278(2):R476-485.

ce
156. Schneider JE, Buckley CA, Blum RM, Zhou D, Szymanski L, Day DE, Bartness TJ.
Ac
Metabolic signals, hormones and neuropeptides involved in control of energy balance
and reproductive success in hamsters. Eur J Neurosci. 2002;16(3):377-379.
157. Schneider JE, Zhou D. Interactive effects of central leptin and peripheral fuel
oxidation on estrous cyclicity. Am J Physiol. 1999;277(4 Pt 2):R1020-1024.
42
158. Farooqi IS, Jebb SA, Langmack G, Lawrence E, Cheetham CH, Prentice AM, Hughes
IA, McCamish MA, O'Rahilly S. Effects of recombinant leptin therapy in a child with
congenital leptin deficiency. N Engl J Med. 1999;341(12):879-884.
159. Chan JL, Heist K, DePaoli AM, Veldhuis JD, Mantzoros CS. The role of falling leptin
levels in the neuroendocrine and metabolic adaptation to short-term starvation in
healthy men. J Clin Invest. 2003;111(9):1409-1421.
160. Welt CK. Will leptin become the treatment of choice for functional hypothalamic
t
ip
amenorrhea? Nat Clin Pract Endocrinol Metab. 2007;3(8):556-557.
cr
161. Welt CK, Chan JL, Bullen J, Murphy R, Smith P, DePaoli AM, Karalis A, Mantzoros
CS. Recombinant human leptin in women with hypothalamic amenorrhea. N Engl J
Med. 2004;351(10):987-997.
us
an
162. Walczewska A, Yu WH, Karanth S, McCann SM. Estrogen and leptin have
differential effects on FSH and LH release in female rats. Proc Soc Exp Biol Med.
M
1999;222(2):170-177.
163. Crane C, Akhter N, Johnson BW, Iruthayanathan M, Syed F, Kudo A, Zhou YH,
e d
Childs GV. Fasting and glucose effects on pituitary leptin expression: is leptin a local
pt
signal for nutrient status? J Histochem Cytochem. 2007;55(10):1059-1074.
164. Sarmento-Cabral A, Peinado JR, Halliday LC, Malagon MM, Castano JP, Kineman
ce
RD, Luque RM. Adipokines (Leptin, Adiponectin, Resistin) Differentially Regulate

Ac
All Hormonal Cell Types in Primary Anterior Pituitary Cell Cultures from Two
Primate Species. Sci Rep. 2017;7:43537.
165. Childs GV, MacNicol AM, MacNicol MC. Molecular Mechanisms of Pituitary Cell
Plasticity. Front Endocrinol (Lausanne). 2020;11(656).
43
166. Allensworth-James ML, Odle AK, Lim J, LaGasse AN, Miles TK, Hardy LL, Haney
AC, MacNicol MC, MacNicol AM, Childs GV. Metabolic signalling to somatotrophs:
Transcriptional and post-transcriptional mediators. J Neuroendocrinol. 2020:e12883.
167. Odle A, Allensworth-James M, Akhter N, Syed M, Haney A, MacNicol M, MacNicol
A, Childs G. A Sex-Dependent Tropic Role for Leptin In The Somatotrope As A
Regulator of POU1F1 and POU1F1-dependent Hormones. Endocrinology,.
2016;157(10):3958-3971.
t
ip
168. Childs GV. Growth hormone cells as co-gonadotropes: partners in the regulation of
cr
the reproductive system. Trends Endocrinol Metab. 2000;11(5):168-175.
169. Childs GV. Development of gonadotropes may involve cyclic transdifferentiation of
us
growth hormone cells. Archives of physiology and biochemistry. 2002;110(1-2):42-
an
49.
170. Childs GV, Unabia G, Miller BT. Cytochemical detection of gonadotropin-releasing

M
hormone-binding sites on rat pituitary cells with luteinizing hormone, follicle-
stimulating hormone, and growth hormone antigens during diestrous up-regulation.

e d
Endocrinology. 1994;134(4):1943-1951.
pt
171. Childs GV, Unabia G, Miller BT, Collins TJ. Differential expression of gonadotropin
and prolactin antigens by GHRH target cells from male and female rats. J Endocrinol.
ce
1999;162(2):177-187.
Ac
172. Childs GV, Unabia G, Rougeau D. Cells that express luteinizing hormone (LH) and
follicle-stimulating hormone (FSH) beta-subunit messenger ribonucleic acids during
the estrous cycle: the major contributors contain LH beta, FSH beta, and/or growth
hormone. Endocrinology. 1994;134(2):990-997.
44
173. Childs GV, Iruthayanathan M, Akhter N, Unabia G, Whitehead-Johnson B.
Bipotential effects of estrogen on growth hormone synthesis and storage in vitro.
Endocrinology. 2005;146(4):1780-1788.
174. Denef C. Paracrinicity: the story of 30 years of cellular pituitary crosstalk. Journal of
175. Odle AK, Allensworth-James M, Haney A, Akhter N, Syed M, Childs GV. Adipocyte
versus Somatotrope Leptin: Regulation of Metabolic Functions in the Mouse.
t
ip
Endocrinology. 2016:en20151811.
cr
176. Allen SJ, Garcia-Galiano D, Borges BC, Burger LL, Boehm U, Elias CF. Leptin
receptor null mice with reexpression of LepR in GnRHR expressing cells display
us
elevated FSH levels but remain in a prepubertal state. American journal of physiology
an
Regulatory, integrative and comparative physiology. 2016;310(11):R1258-1266.
177. Fox RG, Park FD, Koechlein CS, Kritzik M, Reya T. Musashi signaling in stem cells
M
and cancer. Annu Rev Cell Dev Biol. 2015;31:249-267.
178. Szabat M, Kalynyak TB, Lim GE, Chu KY, Yang YH, Asadi A, Gage BK, Ao Z,
e d
Warnock GL, Piret JM, Kieffer TJ, Johnson JD. Musashi expression in beta-cells
pt
coordinates insulin expression, apoptosis and proliferation in response to endoplasmic
reticulum stress in diabetes. Cell Death Dis. 2011;2:e232.

ce
179. Cioffi JA, Shafer AW, Zupancic TJ, Smith-Gbur J, Mikhail A, Platika D, Snodgrass
Ac
HR. Novel B219/OB receptor isoforms: possible role of leptin in hematopoiesis and
reproduction. Nat Med. 1996;2(5):585-589.
180. Brzechffa PR, Jakimiuk AJ, Agarwal SK, Weitsman SR, Buyalos RP, Magoffin DA.
Serum immunoreactive leptin concentrations in women with polycystic ovary
syndrome. J Clin Endocrinol Metab. 1996;81(11):4166-4169.
45
181. Spicer LJ, Francisco CC. The adipose obese gene product, leptin: evidence of a direct
inhibitory role in ovarian function. Endocrinology. 1997;138(8):3374-3379.
182. Chapman IM, Wittert GA, Norman RJ. Circulating leptin concentrations in polycystic
ovary syndrome: relation to anthropometric and metabolic parameters. Clin
Endocrinol (Oxf). 1997;46(2):175-181.
183. Mantzoros CS, Dunaif A, Flier JS. Leptin concentrations in the polycystic ovary
syndrome. J Clin Endocrinol Metab. 1997;82(6):1687-1691.
t
ip
184. Laughlin GA, Morales AJ, Yen SS. Serum leptin levels in women with polycystic
cr
ovary syndrome: the role of insulin resistance/hyperinsulinemia. J Clin Endocrinol
Metab. 1997;82(6):1692-1696.
185.
us
Rouru J, Anttila L, Koskinen P, Penttila TA, Irjala K, Huupponen R, Koulu M. Serum
an
leptin concentrations in women with polycystic ovary syndrome. J Clin Endocrinol
Metab. 1997;82(6):1697-1700.
M
186. Karlsson C, Lindell K, Svensson E, Bergh C, Lind P, Billig H, Carlsson LM, Carlsson
B. Expression of functional leptin receptors in the human ovary. J Clin Endocrinol

e d
Metab. 1997;82(12):4144-4148.
pt
187. Agarwal SK, Vogel K, Weitsman SR, Magoffin DA. Leptin antagonizes the insulin-
like growth factor-I augmentation of steroidogenesis in granulosa and theca cells of

ce
the human ovary. J Clin Endocrinol Metab. 1999;84(3):1072-1076.

Ac
188. Zachow RJ, Magoffin DA. Direct intraovarian effects of leptin: impairment of the
synergistic action of insulin-like growth factor-I on follicle-stimulating hormone-
dependent estradiol-17 beta production by rat ovarian granulosa cells. Endocrinology.
1997;138(2):847-850.
189. Zachow RJ, Weitsman SR, Magoffin DA. Leptin impairs the synergistic stimulation
by transforming growth factor-beta of follicle-stimulating hormone-dependent
46
aromatase activity and messenger ribonucleic acid expression in rat ovarian granulosa
cells. Biol Reprod. 1999;61(4):1104-1109.
190. Spicer LJ, Francisco CC. Adipose obese gene product, leptin, inhibits bovine ovarian
thecal cell steroidogenesis. Biol Reprod. 1998;58(1):207-212.
191. Kikuchi N, Andoh K, Abe Y, Yamada K, Mizunuma H, Ibuki Y. Inhibitory action of
leptin on early follicular growth differs in immature and adult female mice. Biol
Reprod. 2001;65(1):66-71.
t
ip
192. Duggal PS, Van Der Hoek KH, Milner CR, Ryan NK, Armstrong DT, Magoffin DA,
cr
Norman RJ. The in vivo and in vitro effects of exogenous leptin on ovulation in the
rat. Endocrinology. 2000;141(6):1971-1976.
193.
us
Brannian JD, Hansen KA. Leptin and ovarian folliculogenesis: implications for
an
ovulation induction and ART outcomes. Semin Reprod Med. 2002;20(2):103-112.
194. Ricci AG, Di Yorio MP, Faletti AG. Inhibitory effect of leptin on the rat ovary during
M
the ovulatory process. Reproduction. 2006;132(5):771-780.
195. Almog B, Gold R, Tajima K, Dantes A, Salim K, Rubinstein M, Barkan D, Homburg

e d
R, Lessing JB, Nevo N, Gertler A, Amsterdam A. Leptin attenuates follicular

pt
apoptosis and accelerates the onset of puberty in immature rats. Mol Cell Endocrinol.
2001;183(1-2):179-191.
ce
196. Ruiz-Cortes ZT, Martel-Kennes Y, Gevry NY, Downey BR, Palin MF, Murphy BD.
Ac
Biphasic effects of leptin in porcine granulosa cells. Biol Reprod. 2003;68(3):789-
796.
197. Swain JE, Dunn RL, McConnell D, Gonzalez-Martinez J, Smith GD. Direct effects of
leptin on mouse reproductive function: regulation of follicular, oocyte, and embryo
development. Biol Reprod. 2004;71(5):1446-1452.
47
198. Ryan NK, Woodhouse CM, Van der Hoek KH, Gilchrist RB, Armstrong DT, Norman
RJ. Expression of leptin and its receptor in the murine ovary: possible role in the
regulation of oocyte maturation. Biol Reprod. 2002;66(5):1548-1554.
199. Paula-Lopes FF, Boelhauve M, Habermann FA, Sinowatz F, Wolf E. Leptin promotes
meiotic progression and developmental capacity of bovine oocytes via cumulus cell-
independent and -dependent mechanisms. Biol Reprod. 2007;76(3):532-541.
200. Di Yorio MP, Bilbao MG, Pustovrh MC, Prestifilippo JP, Faletti AG. Leptin
t
ip
modulates the expression of its receptors in the hypothalamic-pituitary-ovarian axis in
cr
a differential way. J Endocrinol. 2008;198(2):355-366.
201. Di Yorio MP, Bilbao MG, Biagini-Majorel AM, Faletti AG. Ovarian signalling
us
pathways regulated by leptin during the ovulatory process. Reproduction.
an
2013;146(6):647-658.
202. Karamouti M, Kollia P, Kallitsaris A, Vamvakopoulos N, Kollios G, Messinis IE.

M
Modulating effect of leptin on basal and follicle stimulating hormone stimulated
steroidogenesis in cultured human lutein granulosa cells. J Endocrinol Invest.

e d
2009;32(5):415-419.
pt
203. Ye Y, Kawamura K, Sasaki M, Kawamura N, Groenen P, Sollewijn Gelpke MD,
Kumagai J, Fukuda J, Tanaka T. Leptin and ObRa/MEK signalling in mouse oocyte

ce
maturation and preimplantation embryo development. Reprod Biomed Online.

Ac
2009;19(2):181-190.
204. Tena-Sempere M, Manna PR, Zhang FP, Pinilla L, Gonzalez LC, Dieguez C,
Huhtaniemi I, Aguilar E. Molecular mechanisms of leptin action in adult rat testis:
potential targets for leptin-induced inhibition of steroidogenesis and pattern of leptin
receptor messenger ribonucleic acid expression. J Endocrinol. 2001;170(2):413-423.
48
205. Tena-Sempere M, Pinilla L, Zhang FP, Gonzalez LC, Huhtaniemi I, Casanueva FF,
Dieguez C, Aguilar E. Developmental and hormonal regulation of leptin receptor
(Ob-R) messenger ribonucleic acid expression in rat testis. Biol Reprod.
2001;64(2):634-643.
206. Hoggard N, Mercer JG, Rayner DV, Moar K, Trayhurn P, Williams LM. Localization
of leptin receptor mRNA splice variants in murine peripheral tissues by RT-PCR and
in situ hybridization. Biochem Biophys Res Commun. 1997;232(2):383-387.
t
ip
207. Suter KJ, Pohl CR, Wilson ME. Circulating concentrations of nocturnal leptin, growth
cr
hormone, and insulin-like growth factor-I increase before the onset of puberty in
agonadal male monkeys: potential signals for the initiation of puberty. J Clin
Endocrinol Metab. 2000;85(2):808-814.

us
an
208. El-Hefnawy T, Ioffe S, Dym M. Expression of the leptin receptor during germ cell
development in the mouse testis. Endocrinology. 2000;141(7):2624-2630.

M
209. Zhang J, Gong M. Review of the role of leptin in the regulation of male reproductive
function. Andrologia. 2018.

e d
210. Tena-Sempere M, Barreiro ML. Leptin in male reproduction: the testis paradigm. Mol
pt
Cell Endocrinol. 2002;188(1-2):9-13.
211. Tena-Sempere M, Pinilla L, Gonzalez LC, Dieguez C, Casanueva FF, Aguilar E.

ce
Leptin inhibits testosterone secretion from adult rat testis in vitro. J Endocrinol.
Ac
1999;161(2):211-218.
212. Caprio M, Isidori AM, Carta AR, Moretti C, Dufau ML, Fabbri A. Expression of
functional leptin receptors in rodent Leydig cells. Endocrinology. 1999;140(11):4939-
4947.
49
213. Isidori AM, Caprio M, Strollo F, Moretti C, Frajese G, Isidori A, Fabbri A. Leptin
and androgens in male obesity: evidence for leptin contribution to reduced androgen
levels. J Clin Endocrinol Metab. 1999;84(10):3673-3680.
214. Banks WA, McLay RN, Kastin AJ, Sarmiento U, Scully S. Passage of leptin across
the blood-testis barrier. Am J Physiol. 1999;276(6):E1099-1104.
215. Wang X, Zhang X, Hu L, Li H. Exogenous leptin affects sperm parameters and
impairs blood testis barrier integrity in adult male mice. Reprod Biol Endocrinol.
t
ip
2018;16(1):55.
cr
216. Martins AD, Moreira AC, Sa R, Monteiro MP, Sousa M, Carvalho RA, Silva BM,
Oliveira PF, Alves MG. Leptin modulates human Sertoli cells acetate production and
us
glycolytic profile: a novel mechanism of obesity-induced male infertility? Biochim
an
Biophys Acta. 2015;1852(9):1824-1832.
M
e d
pt
ce
Ac
50
Figure legends
Figure 1. Serum leptin levels in different physiological states. Figure 1A. Serum leptin in mice in
which leptin was ablated in somatotropes (Cre-Gh X Lep fl/fl or adipocytes (Cre-Adipoq X Lepfl/fl).
Controls for each group are littermates from each line, which bore only floxed leptin (Lepfl/fl). In both
females and males, only mice lacking Leptin in adipocytes showed a significant loss in serum leptin
(*=significantly lower than all other values, ANOVA followed by Tukey's Posthoc test). Method:
Serum leptin was assayed by EIA (R&D Systems, Quantikine, MOB00). Figure 1B. Sera were
t
ip
collected from male and female FVB.129P mice during neonatal development and assayed for leptin
as in Figure 1A. In both males and females, the peak levels were seen on day 10. Analysis by Two
cr
Way ANOVA showed no significant sex differences and a significant postnatal age variance: F =
us
14.44. DFn = 6, DFd = 64, P< 0.0001. Tukey's posthoc test identified significant differences. *=data
points that are different from day 1 values. Day 10 values are higher than all others p<0.0001) n/age group
an
ranges from 3--15 mice.
M
Figure 2. Pathways for leptin's permissive actions in the hypothalamus. Stimulation by leptin is
d
mediated by neurons in the ventral premammillary nucleus (PMV), which send connections to
e
Kisspeptin neurons in the anteroventral periventricular and caudal arcuate nuclei and stimulate Kiss1
pt
expression by glutamate, PACAP or nitric oxide. Also, leptin normally inhibits the orexigenic
ce
AgRP-NPY-GABAergic neurons in the Arcuate. When leptin signals are reduced (by food
deprivation, for example), these neurons signal Kisspeptin neurons in the AVPV and caudal Arcuate,
Ac
inhibiting the expression of Kiss1 by AgRP or GABA. These actions ultimately stimulate or inhibit
GnRH neurons and modulate pulsatile activity to effect gonadotropin secretion and the LH surge.
51
Figure 3. Leptin's Role in the Anterior Pituitary. Leptin receptors on gonadotropes and
somatotropes receive rising serum leptin which peaks at midcycle. Leptin stimulates gonadotropes to
produce GnRHR proteins and Fshb and activin mRNAs in partnership with pulsatile GnRH and
estradiol positive feedback. Leptin also stimulates the translation of GH and GHRHR proteins to
support the co-gonadotropic function of somatotropes. The products in green are the known leptin
targets.
t
ip
Figure 4. Leptin's Role in Ovarian Follicular and Oocyte Maturation. All ovarian follicular cells
cr
have leptin receptors and physiological levels of leptin stimulate granulosa and theca cells and oocyte
maturation. This drawing is based on many studies showing leptin stimulation of these target cells.
us
Physiological levels of leptin stimulates, in partnership with growth factors (IGF-1), GH and FSH, to
an
promote the development of follicles to the antral stage. Leptin also promotes ovulation and oocyte
maturation in partnership with LH. Leptin levels rise in association with rising estradiol from the
M
follicles (granulosa cells). Leptin also acts with LH and growth factors to promote oocyte meiosis
and formation of the polar body.

e d
pt
ce
Ac
52
wnloaded from https://academic.oup.com/endo/advance-article/doi/10.1210/endocr/bqaa204/5962079 by San Francisco State University Library user on 14 November 20
ip
cr
us
an
M
ed
pt
ce
Ac
54
ip
cr
us
an
M
ed
pt
ce
Ac
55
ip
cr
us
an
M
ed
pt
ce
Ac
56
ip
cr
us
an
M
ed
pt
ce
Ac

Childs Et Al., 2021 ING

Uploaded by

Document Information

Original Title

Copyright

Available Formats

Share this document

Share or Embed Document

Sharing Options

Did you find this document useful?

Is this content inappropriate?

Copyright:

Available Formats

Childs Et Al., 2021 ING

Uploaded by

Copyright:

Available Formats

The Importance of Leptin to Reproduction

Department of Neurobiology and Developmental Sciences,

University of Arkansas for Medical Sciences, Little Rock, AR 72205

The authors have nothing to disclose

(Kiss1) neurons to stimulate gonadotropin releasing hormone neurons (GnRH). Suppression of

to decreased hypothalamic Gonadotropin-releasing hormone (GnRH) stimulation (7-10). When

In order to respond to nutritional challenges, metabolic signals relay information on energy

dynamic role in reproduction (17-27).

History of the discovery of leptin. Researchers sought answers to questions related to

their HPG axis was immature(31).

Regulation of leptin secretion from adipocytes. As reviewed by Cammisotto et al (38), studies

adipocytes may be stimulated by dexamethasone and other glucocorticoids and peroxisome

proliferator-activated receptor-gamma agonists. Leptin is inhibited by catecholamines, free fatty

by nutrient signals, including glycolytic substrates (glucose, fructose or pyruvate). In addition,

Control of Leptin Signaling

may transport leptin, modulating its bioavailability(47).

(MAPK) (57,58); Phosphoinositol 3 kinase (PI3K) (59,60), mammalian Target of rapamycin

leptin actions and attenuation of leptin resistance in models of diet-induced obesity(47,48).

metabolic and reproductive function in the adult.

by maternal undernutrition (82,84) or administration of a leptin antagonist(90), with diverse

reproductive and metabolic health as the level.

initially implicated as a positive regulator of puberty, because injections of leptin in weanling

amplitude as mechanisms leptin uses to permit development of adult reproductive competence.

reduction in serum leptin in males (109).

Leptin Interactions with Hypothalamic-Pituitary-Gonadal Target Cells

stimulation or suppression of the reproductive system.

Tests of responses to exogenous leptin showed a blunted LH response. However, exogenous

they showed PACAP stimulation of a subset of these neurons(123).

in mice lacking leptin (ob/ob mice) (127,128).

gonadotropes in leptin-deficient mice (135,138-140).

gonadotropins in vivo (31,97,115,143,145-149). For example, leptin restored LH secretion in fasted

as direct metabolic sensors has been a subject of controversy. Are gonadotropes

infertility and abnormal estrous cycling (98).

the leptin influence on somatotropes as well.

multipotential Pou1f1/Pit1 pituitary precursor cell population that perhaps become

by restoring it in mice genetically engineered to be ubiquitously deficient in LEPR (176). They

FSH as a downstream leptin target (97).

functional role had been determined.

regulate differentiated gonadotropes by mediating their expression of GnRHR. Collectively, these

resulting in an activated gonadotrope population(98).

Similarly, in mice, lower concentrations of leptin increased LH or FSH-stimulated ovarian steroids

resistance as a result of exposure to high concentrations.

impaired with inhibitors of either the JAK/STAT or MAPK pathways (202).

stimulatory at the level of the hypothalamus and pituitary (23,64,106,149,153,154,207,209,210).

inhibits testicular function if serum levels rise above a certain threshold.

demonstrated that obesity levels of leptin inhibited gonadotropin-mediated testosterone secretion

(23,210-213). The leptin suppression of testosterone was accompanied by parallel reductions in

Integrating Leptin Actions along the HPG Axis.

results in synchronized leptin-LH pulses and ovulation.

development for the next cycle.

during the writing of this review.

the rat. J Physiol. 1963;166:408-418.

food restriction. Physiol Behav. 1997;61(3):387-394.

female mice and male rats. Am J Physiol. 1986;250(3 Pt 2):R370-376.

8. Cameron JL, Nosbisch C. Suppression of pulsatile luteinizing hormone and

monkey (Macaca mulatta). Endocrinology. 1991;128(3):1532-1540.

pulsatile luteinizing hormone secretion in men after forty-eight hours of fasting. J

Clin Endocrinol Metab. 1991;73(1):35-41.

10. Foster DL, Nagatani S. Physiological perspectives on leptin as a regulator of

reproduction: role in timing puberty. Biol Reprod. 1999;60(2):205-215.

concentration independently estimate percentage body fat in older adults. Am J Clin

Steiner RA. Leptin is a metabolic signal to the reproductive system. Endocrinology.

puberty in normal female mice. J Clin Invest. 1997;99(3):391-395.