SURVEY OF OPHTHALMOLOGY VOLUME 23. NUMBER 6 ??

MAY-JUNE 1979

Experimental Studies on the

Mechanism of Action of Timolol

ARTHUR H. NEUFELD, Ph.D.

Eye Research Institute of the Retina Foundation, and Department of Ophthalmoìogy,

Harvard Medical School, Boston, Massachusetts

Abstract. Timolol, which binds to beta-adrenergic receptors, is a potent antagonist

of the catecholamine-stimulated synthesis of cyclic AMP. However, the actual
mechanism of action by which timolol reduces intraocular pressure is not readily ap-
parent. Compared to its efficacy in human eyes, the drug is relatively ineffective in
rabbit eyes. A reasonable postulate is that soon after administration, timolol blocks
endogenous adrenergic stimulation contributing to the formation of aqueous humor
by the ciliary processes. Nevertheless, the long-lasting reduction of intraocular
pressure persists at a time when the drug is no longer bound to beta-adrenergic recep-
tors. (Surv Ophthalmol 23:363-370, 1979)

Key words. beta adrenergic receptor ?? catecholamines ?? cyclic AMP

- intraocular pressure ?? timolol

P
harmacological manipulation of the
adrenergic nervous system of the eye has
proven to be useful in controlling intra-
ocular pressure. Unlike other physiological
phenomena in which alpha- and beta-adrenergic
stimulation have opposite influences that are
blocked by antagonists, intraocular pressure
decreases after administration of a wide
variety of adrenergic drugs. For example,
therapeutic control of elevated intraocular
pressure can be achieved by applying either a
beta-adrenergic agonist or antagonist directly
to the eye. Although the same response to
pharmacologically opposite drugs might at
first seem perplexing, it is not unreasonable
considering the multicompartmentalized
phenomena associated with the inflow and the
outflow of aqueous humor. Thus, the
363
response to any one adrenergic drug is the
algebraic sum of the responses to stimulation
or inhibition of many adrenergic receptors
with varying sensitivities in different cellular
loei (vascular, epithelial and endothelial)
which al1 contribute to the regulation of
aqueous humor dynamics.
The beta-adrenergic antagonist, timolol, is
now available for use in the treatment of
glaucoma. It is, therefore, appropriate to
review the laboratory investigations which
have contributed to our knowledge concem-
ing the mechanism by which this drug reduces
intraocular pressure. Before doing this, how-
ever, we wil1 review, briefly, adrenergic phys-
iology and pharmacology and, in somewhat
more detail, the relationship between
catecholamines, beta-adrenergic receptors
364 Surv Ophthalmol 23 (6) May-June 1979
and adenosine 3’,5’-monophosphate (cyclic
AMP). The interested reader is referred to
the pertinent chapters in The Phar-
macological Basis of Therapeutics,7 by L. S.
Goodman and A. Gilman or several recent
monographs.2J9J1
Adrenergic Physiology and Pharmacology
Catecholamines are delivered to their tar-
get tissues in the eye via two routes. Nor-
epinephrine is the major neurotransmitter
stored in dense core vesicles in adrenergic
nerve terminals and, upon neural stimulation,
is released into the junctional region between
nerve and responsive cell. Epinephrine is
primarily delivered via the circulation.
Although the eye is densely innervated by the
adrenergic nervous system, the action of
epinephrine as a humoral mediator has not
been clearly demonstrated. Fig. 1 is therefore
drawn to emphasize neuronal input to ocular
tissue and reference should be made to this
figure for the following discussion.
Catecholamines, arriving at the target
tissues do not enter the cells to accomplish
stimulation, but bind to receptors on the cel1
membrane. These receptors are defined,
pharmacologically, by their affinities for test
compounds. The alpha-adrenergic receptor
has a higher affinity for phenylephrine and
norepinephrine than for isoproterenol,
whereas the beta-adrenergic receptor has a
higher affinity for isoproterenol than for
norepinephrine or phenylephrine. Both recep-
tors have high affinities for epinephrine.
Physiologically, the naturally occurring
catecholamines, norepinephrine and epi-
nephrine, bind to both alpha- and beta-
adrenergic receptors on the responsive cel1
membranes. In adrenergically innervated
smooth muscle, such as the iris dilator or
blood vessels, stimulation of alpha-adrenergic
receptors initiates contraction or vaso-
constriction, whereas stimulation of beta-
adrenergic receptors promotes relaxation or
vasodilation. In many nonsmooth muscle
tissues such as liver, fat, and secreting
epithelia, the major influence of epinephrine
is at beta-adrenergic receptors.
Termination of catecholaminergic stimula-
tion is accomplished by uptake of the
catecholamines into the surrounding cells,
thereby effectively decreasing the concentra-
tion of the transmitter at the receptors. This is
NEUFELD
the major route of inactivation of catechol-
amines and two independent uptake pathways
are available. Uptake 1 is into the nerve ter-
minal and is more specific for norepinephrine
than for epinephrine and catecholamine
derivatives. Intracellularly, much of the
catecholamine is degraded, primarily by
monoamine oxidase but also by catechol-O-
methyl transferase. Some catecholamine is
repackaged into the vesicle and reused as
neurotransmitter. Uptake 11 is into ex-
traneuronal cells, i.e., the postjunctional
responsive cells and is nonspecific, taking up
catecholamines and their derivatives. Once
internalized, the catecholamine is metab-
olized, primarily by catechol-0-methyl trans-
ferase and, to a lesser extent, by monoamine
oxidase.
Pharmacological manipulation of the
adrenergic nervous system can be ac-
complished at many levels. Direct acting
agonists, such as norepinephrine, epineph-
rine, isoproterenol or other available deriv-
atives can be exogenously administered to the
eye in high concentrations to ensure penetra-
tion and access to receptor sites. Drugs may
activate alpha-adrenergic receptors, beta-
adrenergic receptors, or have agonistic ac-
tivity at both receptors. A new approach has
used the prodrug, dipivalyl epinephrine. The
enhanced permeability of this lipophilic
derivative of epinephrine allows its use at con-
centrations lower than native catecholamine.
As the compound penetrates the eye, the
dipivalyl moieties are removed by enzymatic
hydrolysis, liberating epinephrine for direct
action at the receptors.
Endogenous influences on a tissue may be
altered or blocked by antagonistic drugs
which bind to specific receptors for
catecholamines. Compounds such as the
alpha-adrenergic antagonist, thymoxamine,
and the beta-adrenergic antagonists,
propranolol, timolol, and atenolol, have been
used recently to accomplish adrenergic recep-
tor blockade in anterior segment tissues. We
emphasize, however, that for phar-
macological antagonists to be active, there
must be tonic stimulation which they black or
an opposing physiological influence which
prevails.
Indirect acting, sympathomimetic drugs
such as tyramine, amphetamine, reserpine,
and guanethadine cause the release of
catecholamine from nerve terminals and
reduce the inactivation and storage capability
MECHANISM OF ACTION OF TIM0101 365

FIG. I. Adrenergic neuromuscular junction: hypothetical action of timolol at the /3-adrenergic receptor.
Timolol, T; epinephrine, E; norepinephrine, NE; cr-adrenergic receptor, (Y; /3-adrenergic receptor, @;
cyclic AMP, CAMP; stimulatory, +; inhibitory, -; neuronal uptake, I, extraneuronal uptake, II;
monoamine oxidase, MAO, catechol-O-methyl transferase, COMT. (Peter Mallen, artist)

of the tissue. Under these conditions, en-
dogenously available or exogenously ad-
ministered catecholamines have an enhanced
potency because the termination pathway has
been compromised and the effective concen-
tration of agonist at the receptors is thereby
maintained for greater periods of time. Alter-
native methods to accomplish this type of
potentiation include direct inhibition of
neuronal uptake, accomplished phar-
macologically with cocaine, protriptyline or
imipramine. Also, local injection of 6-
hydroxydopamine, which chemically causes
the destruction of the nerve terminals,
produces a sustained elimination of Uptake I.
Additional pharmacological approaches to
maintain effective concentrations of
catecholamine at the receptor involve inhibi-
tion of metabolism by monoamine oxidase
and catechol-O-methyl transferase. Drugs
such as pargyline, nialamide, tranylcypro-
mine and pyrogallol inhibit one or the other
of these enzymes, thereby interfering with
metabolic inactivation. However, because
this step occurs after uptake has removed the
catecholamine from the vicinity of the recep-
tor, blocking metabolism does not substan-
tially potentiate the action of agonists.
Beta-Adrenergic Receptors
Epinephrine, arriving at target cells via the
circulation, or norepinephrine, arriving at
target cells via the innervation, binds to beta-
adrenergic receptors. These receptors are on
366 Surv Ophtholmol 23 (6) May-June 1979
the cell surface and diffuse laterally in the
membrane, retaining an orientation that
allows interaction with extracellular
catecholamines. Apparently, the occupied,
rapidly diffusing receptors have a greater
ability to activate adenylate cyclase, the en-
zyme that catalyzes the synthesis of cyclic
AMP, than do the unoccupied receptors;
probably, an intramembrane coupling moiety
is involved in the link between receptors and
adenylate cyclase.
The beta-adrenergic receptors are defined
by their affinities which are greater for beta-
adrenergic agonists and antagonists than for
alpha-adrenergic agonists and antagonists.
Thus, beta-adrenergic receptors have high af-
finities for agonists, such as isoproterenol and
epinephrine, and antagonists, such as pro-
pranolol and timolol, are very tightly bound.
The alpha-adrenergic antagonists, phenoxy-
benzamine and phentolamine, do not bind to
the beta-adrenergic receptors. Pharmaco-
logical evidence indicates that there are two
types of beta-adrenergic receptors: the beta,-
adrenergic receptors, which are character-
istic of adipose and cardiac tissue and have a
relatively high affinity for norepinephrine,
and the beta,-adrenergic receptors, which are
characteristic of pulmonary and vascular
smooth muscle and have a relatively low af-
finity for norepinephrine. Practolol, atenolol
and sotalol selectively bind to betal-
adrenergic receptors and salbutamol and
butoxamine selectively bind to betaz-
adrenergic receptors. Propranolol and
timolol bind to both types of receptors. For
the iris-ciliary body of rabbits, the relative
abilities of isoproterenol, epinephrine and
norepinephrine to displace radiolabeled
dihydroalprenolol from membranes prepared
from this tissue indicate a predominance of
beta,-adrenergic receptors.15
Adenylate cyclase is oriented in the mem-
brane so that, when coupling occurs between
it and the occupied beta-adrenergic receptor,
cyclic AMP is formed intracellularly from
ATP. Because many tissues can synthesize
cyclic AMP, the specificity for the response
pathway is provided by the receptor for the
transmitter. When cyclic AMP acts as a sec-
ond messenger, the transduction of the
catecholaminergic signal occurs only at target
cells with beta-adrenergic receptors. Con-
versely, cells lacking beta-adrenergic recep-
tors will not increase their intracellular con-
centrations of cyclic AMP when challenged
NEUFELD
with catecholamines.
Although some basal amount of cyclic
AMP is formed in the absence of stimula-
tion, the increase in the concentration of
cyclic AMP that mediates the physiological
response to catecholamines is due to de nuvo
synthesis: It is not surprising, then, that intra-
cellular increases in cyclic AMP occur
quickly. The activated receptor-adenylate
cyclase complex continues to synthesize cyclic
AMP until the extracellular catecholamine
concentration decreases sufficiently and un-
occupied receptors accumulate. As dissocia-
tion of the receptor-adenylate cyclase com-
plex occurs, the enzyme returns to its basal
level of activity.
Beta-adrenergic antagonists specifically
block the ability of catecholamines to bind to
the beta-adrenergic receptors and, therefore,
activation of adenylate cyclase does not oc-
cur. Although catecholamine-stimulated syn-
thesis of cyclic AMP is prevented by the an-
tagonist, the basal level of cyclic AMP is not
affected nor is the response to other
transmitters. Antagonists are pharmaco-
logical agents and there are no known
physiological antagonists.
Timolol and Intraocular Pressure
Compared to the number of clinical studies
demonstrating the usefulness of timolol, there
have been relatively few laboratory investiga-
tions of the mechanism by which this drug in-
fluences intraocular pressure. Generally, the
work that has been done confirms the clinical
observations that timolol lowers the intra-
ocular pressure of the normotensive eyelo and
is particularly effective in the hypertensive
eye. 1,11,24
In rabbits, although the reponse to topical
timolol is small, it has been reported to be
effective by two groups of investigators.“Q”
As in humans, the time of onset of action of
timolol is short. After topical application, the
drug appears rapidly in the aqueous humor,
reaching a peak approximately thirty minutes
after administration and is detectable in the
plasma even earlier. 2o Within thirty minutes,
timolol significantly decreases intraocular
pressure in the normal rabbit eye.18J0
Vareilles et alZo compared the efficacy of
several adrenergic agents on the intraocular
pressure in rabbits. In normotensive animals,
topical epinephrine and isoproterenol de-
MECHANISM Of ACTION OF JIMOLOL
crease intraocular pressure more effectively
and consistently than propranolol or timolol.
In animals with experimental ocular hyper-
tension produced several months earlier by
pretreating intracamerally with alpha-chymo-
trypsin, 0.5% topical timolol causes a large
decrease in intraocular pressure which is ap-
parent within 60 minutes and is maximal
about three hours later. 1.5% timolol is not
more effective. Whereas the magnitude of the
influence of timolol is similar to that of
isoproterenol and norepinephrine, these latter
compounds have maximal effects about one
hour earlier than timolol. Propranolol is not
as effective as timolol in this model.
Vareilles et a12”and Radius et all6 found no
loss of effectiveness of timolol when given
repeatedly to rabbits, two or three times daily
for a week. Although the duration of action of
timolol is less than sixteen hours, Radius et
alI6 report that, when repeated administra-
tion of the drug is discontinued, the intra-
ocular pressure does not return to pretreat-
ment levels for 48 hours.
When ocular hypertension is induced in
rabbits by water loading, pretreatment with
timoiol prevents, to some extent, the increase
in intraocular pressure which normally occurs
twenty minutes after provocation.20 1.5%
timolol is not more effective and 1.5%
propranolol has similar activity. In this
model, both epinephrine and isoproterenol, at
equivalent dosages, are more effective for a
longer duration than timolol. However,
timolol potently blocks the ability of
isoproterenoi to prevent the increase of intra-
ocular pressure after water loading.
Radius et all6 report that the contralateral
eye, not treated with timolol, also has a
decreased intraocular pressure, often of
similar magnitude to the treated eye. Thus,
one hour after the initial unilateral dose of
timolol, the intraocular pressure in both eyes
decreases by 7-8 mm Hg. The reduction in
the untreated eye persists with repeated daily
treatment and such a contralateral response
has been reported in clinical studies.25
When topical treatment with timolol is
followed one hour later with application of
2% norepinephrine or epinephrine, a further
decrease in intraocular pressure occurs after
several hours and persists longer in eyes
treated with the combination of drugs as com-
pared to treatment with timolol alone. This
additional influence of adrenergic agonists in
the presence of timolol does not occur with
367
albuterol, a beta-adrenergic agonist. Radius
et alI6 conclude that the reduction of intra-
ocular pressure by catecholamines in the
presence of timolol is due to alpha-adrenergic
stimulation which is not blocked by the beta-
adrenergic antagonist. In patients, additional
reduction of intraocular pressure by
epinephrine in eyes treated with timolol has
been observed occasionally,” but not con-
sistently.’
Timolol lowers intraocular pressure by
decreasing the formation of aqueous humor.
Although not demonstrated in laboratory
animals, the results of clinical observations
using tonography indicate that topical timolol
has little or no influence on outflow
facility.‘8,23 The conclusion from these
studies, that the decrease in intraocular
pressure must be due to decreased inflow, was
substantiated recently by more direct
measurements on human volunteers. Using
ffuorophotometry, as originally described by
Jones and Maurice,g Yablonski et a122and
Coakes and Brubake? found that timolol
decreases the turnover rate of aqueous
humor. Previously, Missotten and Goethals”
had demonstrated that topical timoloi
decreases the light-insensitive potential
measured as part of the electrooculogram in
normal adult subjects, which is consistent
with an inhibitory action of the drug on the
secretion of fluid by the ciliary processes.
Cellular Mechanism of Action
We have been interested in studying the
mechanisms by which timolol influences the
normal cellular physiology governing the for-
mation of aqueous humor. Because of the
known beta-adrenergic antagonistic activity
of timolol, we investigated the interaction of
timolol with catecholamines, beta-adrenergic
receptors and the stimulation and inhibition
of the synthesis of cyclic AMP in anterior
segment tissues.
In studies in our laboratory, topically
applied timolol did not increase cyclic AMP
levels in the aqueous humor of rabbit eyes,
nor stimulate cyclic AMP formation by
ocular tissues, indicating that the drug has no
intrinsic beta-adrenergic agonistic activity
(Bartels, Jumblatt, Neufeld, in preparation).
When we compared the ability of adrenergic
agents to displace radiolabeled dihydro-
alprenolol from beta-adrenergic receptors on
368 Surv Ophtholmol 23 (6) May-June 1979
membranes prepared from rabbit iris-ciliary
bodies, we found that timolol is ap-
proximately equipotent with propranolol and
both compounds are more potent than the
agonists: epinephrine, isoproterenol, and
norepinephrine.12 In vitro, timolol blocks the
stimulation by catecholamines to increase
cyclic AMP concentrations in ocular tissues
(Bartels, Jumblatt, Neufeld, in preparation)
and thus has the ability to act as a potent
beta-adrenergic antagonist.
The ability of timolol to alter a physio-
logical process such as the formation of
aqueous humor has an important implica-
tion. For such a drug to be active in vivo,
there must be tonic adrenergic stimulation of
the target tissue. By analogy, topical atropine
would not dilate the pupil if the sphincter
muscle were not under cholinergic tone, op-
posed by adrenergic input to the dilator mus-
cle. The finding that timolol is an effective
ocular hypotensive agent suggests that the
drug blocks adrenergic stimulation which
physiologically increases intraocular pressure
by enhancing inflow. There has not been,
however, a demonstration, in vivo, that beta-
adrenergic stimulation in the ciliary processes
maintains or promotes the formation of
aqueous humor.
A major difficulty in investigating the ac-
tion of timolol is our relatively imprecise un-
derstanding of the mechanisms by which
aqueous humor is formed. Timolol may act
on secretion, ultrafiltration, or both phenom-
ena. The secretion of aqueous humor is
dependent on blood flow to the ciliary
processes. If timolol prevented adrenergically
stimulated vasodilation in the blood vessels
leading to the ciliary processes, a net
vasoconstriction would reduce blood flow and
perfusion area, resulting in decreased active
transport by the ciliary epithelium. In addi-
tion, timofol might decrease secretion by
blocking a stimulatory action of catechol-
amines directly on the secretory cells, the
nonpigmented epithelia. Such a mechanism,
implying that cyclic AMP stimulates ion
transport in these cells, has been observed in
other transporting epithelial tissues and was
suggested by experiments by Cole and
Nagasubramania# on isolated rabbit iris-
ciliary body.
Furthermore, vasoconstriction induced by
timolol in afferent vessels in the ciliary
processes might reduce ultrafiltration if
decreased perfusion pressure lowered the
NEUFELD
driving force, i.e., the pressure difference
between the blood vessels and the posterior
chamber. In addition, Green and Griffin’
reported that catecholamines stimulate
pressure-dependent fluid flow through the
ciliary processes in isolated tissue. Should
such a mechanism be active in vivo, perhaps
timolol blocks this component of ultrafiltra-
tion.
If timolol has a vascular locus of action,
one should observe net vasoconstriction.
However, when timolol is applied topically,
marked changes in ocular vessels are not
grossly apparent and are unlikely to be
sustained for the duration of action of the
drug. Although timolol may act initially on a
particularly sensitive vascular network in the
ciliary processes, we favor the explanation
that the major initial action is directly on the
secreting epithelia to block active transport or
ultrafiltrative mechanisms which participate
in the formation of aqueous humor. We point
out, however, that there is little experimental
basis for this mechanism.
Wherever the initial antagonism occurs,
the prolonged action of timolol occurs at a
time when we are not able to demonstrate
that timolol blocks the synthesis of cyclic
AMP. Three hours after topical administra-
tion of timolol, when the maximal reduction
of intraocular pressure occurs, corneas and
iris-ciliary bodies of rabbit eyes were excised
and incubated in vitro. When challenged with
exogenous catecholamines, corneas from
treated eyes synthesize less cyclic AMP than
do corneas from untreated eyes, indicating
the presence and efficacy of antagonism by
timolol. However, the iris-ciliary body tissue
from treated eyes does not have a markedly
impaired ability to synthesize cyclic AMP,
compared to untreated controls (Bartels,
Jumblatt, Neufeld, in preparation). Thus, the
maximal influence of timolol in decreasing
the formation of aqueous humor occurs at a
time when there is not sufficient drug in the
iris-ciliary body to antagonize the stimula-
tion of the synthesis of cyclic AMP by ex-
ogenous catecholamines. We have recently
observed a similar phenomenon in the cornea.
Several days after repeated treatment with
topical timolol, when we can no longer
demonstrate an inhibition of the synthesis of
cyclic AMP,‘* corneas from eyes treated in
vivo still have an impaired ability to transport
chloride in response to exogenous epineph-
rine.3 These findings suggest that timolol
MECHANISM OF ACTION OF TIMOLOL
induces prolonged changes in the physiology
of transporting epithelial cells which outlast
the presence of the drug in the tissue and the
inhibition of catecholamine-stimulated syn-
thesis of cyclic AMP.
There are additional aspects of the action
of timolol which are puzzling but nevertheless
may contribute to its clinical efficacy.
Although direct comparisons have not been
made, one has the impression that timolol
may cause a greater reduction of the in-
traocular pressure of human eyes than can be
demonstrated with laboratory animals. In ad-
dition, when compared for the ability to
reduce intraocular pressure, timolol is more
potent than propranolol; however, both com-
pounds compete equipotently at the beta-
adrenergic receptor on membrane prepara-
tions from iris-ciliary body12 or cardiac
tissue.’ Finally, there is the finding in both
laboratory18 and clinicalz2J4 studies that
timolol, given unilaterally, lowers intraocular
pressure in the contralateral eye. Because it is
unlikely that sufficient timolol reaches the
contralateral eye via the circulation, there
may be a central, as well as a peripheral, ac-
tion of timolol to lower intraocular pressure.
Although the usefulness of timolol for the
treatment of glaucoma has excited many
ophthalmologists, the cellular mechanism of
action of the drug is largely unknown.
Nevertheless, the conclusion from clinical
studies that timolol suppresses inflow has led
to new insights and hypotheses concerning the
regulation of the formation of aqueous humor
which should stimulate the interests of many
laboratory investigators.
References
Boger WP III, Puliafito CA, Steinert RF, et
al: Long-term experience with timolol
ophthalmic solution in patients with open-
angle glaucoma. Ophthalmology 85:259-267,
1978
Burnstock G, Costa M: Adrenergic Neurons.
New York, John Wiley & Sons, 1975
Candia OA, Podos SM, Neufeld AH:
Modification by timolol of catecholamine
stimulation of chloride transport in isolated
corneas. Invest Ophthalmol Vis Sci (in press)
Chenieux-Guicheney P, Dausse JP, Meyer P,
et al: Inhibition of fH]-dihydroalprenolol
binding to rat cardiac membranes by various
369
beta-blocking agents. Br J Pharmacol 63:
177-182, 1978
5. Coakes RI, Brubaker RF: The mechanism of
timolol in lowering intraocular pressure. Arch
Ophthalmol %:2045-2048, 1978
6. Cole DF, Nagasubramanian S: The effect of
natural and synthetic vasopressins and other
substances on active transport in ciliary
epithelium of the rabbit. Exp Eye Res
13:45-57, 1972
7. Goodman LS, Gilman A (Eds): The Phar-
macological Basis of Therapeutics. New York,
Macmillan Publishing Co., Inc., 1975
8. Green K, Griffin C: Adrenergic effects on the
isolated rabbit ciliary epithelium. Exp Eye Res
27:143-149, 1978
9. Jones RF, Maurice DM: New methods of
measuring the rate of aqueous flow in man
with fluorescein. Exp Eye Res 5:208-220, 1966
10. Katz IM, Hubbard WA, Getson AJ, et al: In-
traocular pressure decrease in normal
volunteers following timolol ophthalmic solu-
tion. Invest Ophthaimol Fk489-492, 1976
11. Missotten L, Goethals M: Timolol reduces the
standing potential of the eye. Ophthalmd Res
9:321-323, 1977
12. Neufeld AH, Page ED: In vitro determination
of the ability of drugs to bind to adrenergic
receptors. Invest Ophthalmol Vis Sci 16:
1118-1124, 1977
13. Neufeld AH, Zawistowski KA, Page ED, et
al: Influences on the density of beta adrenergic
receptors in the cornea and iris-ciliary body of
the rabbit. Invest Ophthalmol Vis Sci 17:
1069-1075, 1978
14. Nielsen NV: Timolol: Hypotensive effect,
used alone and in combination for treatment
of increased intraocular pressure. Acta
Ophthalmol 56:504-509, 1978
15. Page ED, Neufeld AH: Characterization of cr-
and P-adrenergic receptors in membranes
from the rabbit iris before and after develop
ment of supersensitivity. Biochem Pharmacol
27:953-958, 1978
16. Radius R, Diamond G, Pollack I, et al:
Timolol: a new drug for management of
chronic simple glaucoma. Arch Ophthalmol
%:1003-1008, 1978
17. Ritch R, Hargett NA, Podos SM: The effect
of 1.5% timolol maleate on intraocular
pressure. Acta Ophthalmol 56:6-10, 1978
18. Sonntag J, Brindley G, Shields M: Effect of
timolol therapy on outflow facility. Invest
Ophthalmol Vis Sci 17:293-296, 1978
19. Triggle DJ, Triggle CR: Chemical Phar-
macology of the Synapse. New York,
Academic Press, 1976
20. Vareilles P, Silverstone D, Plazonnet B, et al:
Comparison of the effects of timolol and other
adrenergic agents on intraocular pressure in
the rabbit. Invest Ophthalmol Vis Sci
16:987-996, 1977
370 Surv Ophtholmol 23 (6) May-June 1979 NEUFELD

2 I. Williams LT, Letkowitz RJ: Receptor Binding response and duration of action. Arch
Studies in Adrenergic Pharmacology. New Ophthalmol 95605607, 1977
York, Raven Press, 1978
22. Yablonski ME, Zimmerman TJ, Waltman
SR, et al: A fluorophotometric study of the Supported in part by U.S. Public Health Service
effect of topical timolol on aqueous humor grants EY-02367, EY-02360, EY-00785, P3EY-
dynamics. Exp Eye Res 27:135-142, 1978 01784, Biomedical Research Support Grant PHS
23. Zimmerman T, Harbin R, Pett M, et al: 5S07RR05527, and Massachusetts Lions Eye
Timolol and facility of outflow. Invest Research Fund, Inc. Dr. Neufeld is the recipient of
Ophthalmol Vis Sci 16:623-624, 1977 Research Career Development Award EY-00114.
24. Zimmerman T, Kaufman H: Timolol: A beta- Reprint requests should be addressed to Arthur
adrenergic btocking agent for the treatment of H. Neufeld, Ph.D., Eye Research Institute of
glaucoma. Arch Ophthalmol95:601-604, 1977 Retina Foundation, 20 Staniford Street, Boston,
25. Zimmerman T, Kaufman H: Timolol: Dose MA 02114.