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Contents lists available at ScienceDirect

Behavioural Brain Research

journal homepage: www.elsevier.com/locate/bbr

Review

Serotonin and conditioning: Focus on Pavlovian psychostimulant drug

conditioning
Robert J. Carey a,b,∗ , Ernest N. Damianopoulos c
a
Research Service and Development (151), VA Medical Center, 800 Irving Avenue, Syracuse, NY 13210, USA
b
Department of Psychiatry and Graduate School, SUNY Upstate Medical University at Syracuse, Syracuse, NY, USA
c
Research Service and Development (151), VA Medical Center, Room 326, 800 Irving Avenue, Syracuse, NY 13210, USA

h i g h l i g h t s

• A review of the serotonin system and its contribution to behavior.

• A review of conditioning and manipulations of the serotonin system.
• Serotonin selective drugs and psychostimulant conditioning.
• A proposed post-trial methodology to assess psychostimulant conditioning.

a r t i c l e i n f o a b s t r a c t

Article history: Serotonin containing neurons are located in nuclei deep in the brainstem and send axons throughout the
Received 1 April 2014 central nervous system from the spinal cord to the cerebral cortex. The vast scope of these connections
Received in revised form 15 October 2014 and interactions enable serotonin and serotonin analogs to have profound effects upon sensory/motor
Accepted 20 October 2014
processes. In that conditioning represents a neuroplastic process that leads to new sensory/motor con-
Available online xxx
nections, it is apparent that the serotonin system has the potential for a critical role in conditioning. In this
article we review the basics of conditioning as well as the serotonergic system and point up the number of
Keywords:
non-associative ways in which manipulations of serotonin neurotransmission have an impact upon con-
Serotonin
Sensory/motor function
ditioning. We focus upon psychostimulant drug conditioning and review the contribution of drug stimuli
Pavlovian conditioning in the use of serotonin drugs to investigate drug conditioning and the important impact drug stimuli can
Psychostimulant drug conditioning have on conditioning by introducing new sensory stimuli that can create or mask a CS. We also review the
Drug stimuli ways in which experimental manipulations of serotonin can disrupt conditioned behavioral effects but
Post-trial drug treatment not the associative processes in conditioning. In addition, we propose the use of the recently developed
memory re-consolidation model of conditioning as an approach to assess the possible role of serotonin
in associative processes without the complexities of performance effects related to serotonin treatment
induced alterations in sensory/motor systems.
© 2014 Elsevier B.V. All rights reserved.

Contents

1. Conditioning: Pavlovian and instrumental/operant conditioning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00

1.1. Pavlovian conditioning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
1.2. Delay and trace conditioning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
1.3. Fundamentals of Pavlovian conditioning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
1.4. Current simplified conditioning model preparations and differential CNS-site involvement . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00

∗ Corresponding author at: Research Service and Development (151), VA Medical Center, 800 Irving Avenue, Syracuse, NY 13210, USA. Tel.: +1 315 682 9251;
fax: +1 3156829251.
E-mail address: careybdjp@earthlink.net (R.J. Carey).

http://dx.doi.org/10.1016/j.bbr.2014.10.038
0166-4328/© 2014 Elsevier B.V. All rights reserved.

Please cite this article in press as: Carey RJ, Damianopoulos EN. Serotonin and conditioning: Focus on Pavlovian psychostimulant drug
conditioning. Behav Brain Res (2014), http://dx.doi.org/10.1016/j.bbr.2014.10.038
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1.5. Mathematical models of Pavlovian conditioning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00

1.6. Instrumental/operant conditioning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
1.7. Pavlovian versus instrumental conditioning: differences . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
2. Serotonin . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
2.1. Basic neuroanatomy of the Serotonin system . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
2.2. Serotonin and motor function . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
2.3. Serotonin and sensory processing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
3. Psychostimulant drug conditioning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
3.1. Psychostimulant drug conditioning and high abuse liability . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
3.2. Paired/unpaired treatment design as a necessary feature in studies of Pavlovian drug conditioning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
3.3. Critical differences of drug conditioning from simplified standard models of Pavlovian conditioning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
3.4. Paradigms and preparations in the study of Pavlovian drug conditioning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
3.5. Characteristic features of the Pavlovian drug conditioning hyper/hypo-activity model . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
4. Methodological issues in the use of experimental manipulations of serotonin to study psychostimulant drug conditioning . . . . . . . . . . . . . . . . . . . . . 00
4.1. Introduction . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
4.2. Methods for the behavioral analysis of serotonin manipulations: “knockout” preparations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
4.3. Methods for the behavioral analysis of serotonin manipulations: “lesion” preparations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
4.4. Behavioral assessment methods: incentive/reward preparations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
4.5. Behavioral assessment methods: aversive learning preparations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
4.6. Behavioral assessments of transmitter manipulations . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
4.7. Significant test environment variables . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
4.7.1. Novelty/familiarity variables . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
4.7.2. Stimulus processing . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
4.8. Drug stimulus effects on conditioning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
4.9. Drug stimulus effects in testing for conditioned drug treatment effects . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
5. A new model to assess the role of serotonin in conditioning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
5.1. Conditioning and the memory trace . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
5.1.1. Memory trace and consolidation phase . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
5.1.2. Memory trace and reconsolidation phase . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
5.2. Neurotransmitter inhibition and reconsolidation of conditioning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
5.3. The post-trial drug treatment model as a tool to assess specific serotonin receptor subtypes implicated in psychostimulant
conditioning . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00
References . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 00

1. Conditioning: Pavlovian and instrumental/operant 1.3. Fundamentals of Pavlovian conditioning

conditioning
In an extensive and detailed series of conditioning experiments,
1.1. Pavlovian conditioning Pavlov and associates established the basics of this conditioning:
(a) the order of the CS–UCS pairing is critical: conditioning occurs
Conditioning is an associational process that induces neuro- only if CS precedes UCS (forward conditioning) but no conditioning
plasticity. The most basic form of conditioning was identified and occurs if UCS precedes and terminates before the CS (backward con-
systematically analyzed by Pavlov [1,2]. This Pavlovian or classical ditioning), (b) following conditioning parametric changes in the CS
conditioning entails the pairing of two qualitatively different stim- result in comparable parametric decrements in the efficacy of the
uli in close temporal contiguity so that after repeated pairings the CS to evoke the CR (stimulus generalization) and (c) after repeated
two stimuli come to evoke the same behavioral response. One of presentations of the CS without the UCS, the CS loses its capacity
the stimuli termed the unconditioned stimulus (UCS) reliably elic- to elicit the CR (extinction). Pavlov also showed that extinction of
its by innate mechanisms a readily identifiable response labeled as the CS–CR connection was not permanent in that after a sustained
the unconditioned response (UCR) and, the other stimulus labeled period of non-exposure to the CS there is a spontaneous recovery
as the conditioned stimulus (CS) is neutral with respect to this UCR. of the capacity of the CS to elicit the CR. This latter observation indi-
It is only after the repeated pairings of the two stimuli that the CS cates that conditioning can induce a relatively permanent change
comes to elicit a response (the CR) that is qualitatively similar but in the nervous system.
not necessarily identical to the UCR previously elicited only by the
UCS.
1.4. Current simplified conditioning model preparations and
differential CNS-site involvement
1.2. Delay and trace conditioning
While Pavlov’s extensive body of experimental work provided
There are various ways in which the CS and UCS can be paired. the basic principles of conditioning, new behavioral and concep-
If the CS is presented such that it overlaps with the UCS it is termed tual models of this conditioning process have been developed. One
a delayed conditioning protocol. In this arrangement the CS and major direction has been in the use of a UCS with a brief onset
UCS occur in temporal contiguity. In this case, the associative rela- and offset of the UCR such as an air puff to the eye to evoke the
tionship is straightforward. Pavlov also showed that conditioning brief reflex response of an eye blink. Using a UCS with a brief onset
could occur if the CS terminated a second or two before the UCS was and offset of the UCR allowed the onset and offset of the UCR and
presented. This was termed trace conditioning. Pavlov assumed in CR to be measured with precision. Indeed, this has enabled delay
this second case that a trace or a CNS representation of the CS had and trace conditioning to be studied in ways that can be tied to
to persist long enough to overlap with the UCS so that temporal brain systems. It is now accepted that delay conditioning with the
contiguity was preserved sufficient to form an association. use of punctate stimuli as conditioned stimuli is associated with

Please cite this article in press as: Carey RJ, Damianopoulos EN. Serotonin and conditioning: Focus on Pavlovian psychostimulant drug
conditioning. Behav Brain Res (2014), http://dx.doi.org/10.1016/j.bbr.2014.10.038
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cerebellar mechanisms [3] whereas the trace conditioning involves
the frontal cortex and hippocampus [4].
1.5. Mathematical models of Pavlovian conditioning
In addition to these detailed empirical investigations tying con-
ditioning with discrete stimuli to brain mechanisms there have
been a number of mathematical modeling efforts to better char-
acterize Pavlovian conditioning e.g., Rescorla and Wagner [5],
Mackintosh [6], Frey and Sears [7], Pearce and Hall [8]. The lack
of relevance of these models to CNS processes has been detailed in
the seminal papers by Damianopoulos [9,10].
1.6. Instrumental/operant conditioning
Another basic type of conditioning is termed instrumental or
trial and error learning. In this type of learning, an organism ini-
tiates the behavioral responses that enable it to obtain a positive
reinforcement such as food or avoid an aversive event such as a
threatening or painful stimulus. Whereas Pavlovian conditioning
is considered involuntary in that the UCS elicits an involuntary
reflexive response that is imposed on the organism, instrumen-
tal conditioning is considered voluntary in that the organism must
first emit the response for the UCS (positive or negative stimuli)
to occur. As important as these distinctions are, in many learning
situations there is a composite of both conditioning processes. In
fact “pure” classical conditioning of autonomic system responses
is difficult to arrange in that the animal typically needs to be
restrained. In Pavlov’s conditioning studies the dogs used for the
conditioned salivation and gastric secretion experiments required
physical restraint to permit the measurement of the autonomic
system responses. If the dogs were not restrained, then, vigorous
behavioral activation in response to the CS would have occurred
and interfered with the collection methods available at that time.
While Pavlov obviously recognized the intense behavioral activa-
tion elicited by the conditioned food stimulus, his focus was upon
the autonomic nervous system response in that the autonomic
nervous system was considered to be immune from instrumen-
tal/operant associative process effects. Indeed, this connection of
Pavlovian conditioning to the autonomic nervous system has been
applied widely to human and animal behavior particularly Psy-
chosomatic Medicine and Drug Addiction. Essentially, what this
conditioning does is to attach an innate autonomic response mech-
anism to an arbitrary unrelated external stimulus so it is easy to
see how this would be of substantial importance to Psychiatry and
Psychology.
1.7. Pavlovian versus instrumental conditioning: differences
The key difference between instrumental and classical or
Pavlovian conditioning is not defined by the nature of the
elicited/emitted response (i.e., autonomic versus skeletal-muscle
motor responses). The difference is in the order and sequence in
which the target response is elicited/emitted in relation to the
UCS. In classical conditioning the response is directly evoked by
the UCS and is not determined by the organism’s ongoing behav-
ior. In instrumental conditioning the response is dependent upon
ongoing behavior in that the selected target response (the IR) first
needs to be emitted by the organism in order to have the UCS pre-
sented; that is, the response is instrumental to the presentation
of the UCS. In instrumental conditioning the response is necessary
for food delivery in a food deprived animal and needs to be emit-
ted by the animal to contingently result in the delivery of the UCS
(i.e., food), whereas, in Pavlovian conditioning the UCS (e.g., food)
is delivered independent of the behavior of the animal.
Please cite this article in press as: Carey RJ, Damianopoulos EN. Serotonin and conditioning: Focus on Pavlovian psychostimulant drug
conditioning. Behav Brain Res (2014), http://dx.doi.org/10.1016/j.bbr.2014.10.038
3
2. Serotonin
2.1. Basic neuroanatomy of the Serotonin system
Serotonin containing neurons comprise a phylogenetically
ancient neurotransmitter network. Serotonin (5-HT) neurons in
the mammalian brain are located in clusters of raphe nuclei that
encompass areas from the midbrain to the medulla. While the total
number of 5-HT neurons is small relative to total neuronal cells
in the brain (e.g., approximately 20 K in rat brain), the axons of
these cells have contacts (primarily paracrine) with many other
neurons so that virtually the entire brain and spinal cord receives
5-HT innervation [11,12]. Consistent with the rostral/caudal dis-
tribution of 5-HT raphe nuclei, the caudal pontine and medullary
clusters send projections to the cerebellum, medulla and spinal
cord, whereas the midbrain raphe nuclei innervate the forebrain.
In keeping with the descending projections to the medulla, the 5-
HT neurons have a substantial impact on basic functions related to
respiration, thermal regulation and cardiovascular function. Pro-
jections from 5-HT nucleus raphe obscurus and ventral lateral
medullary neurons extend to the ventral horn of the spinal cord
and profusely innervate motor neurons to impact motoric function
[12].
More elusive in terms of function are the forebrain projections of
the dorsal and median raphe nuclei situated in the midbrain reticu-
lar formation. These 5-HT axons project to neo- and paleocortex as
well as subcortical striatal and limbic nuclei [13]. Not surprisingly,
perturbations in the forebrain 5-HT have been implicated in several
diverse functions, including cognition, memory, aggression, feed-
ing and reproduction [14–18]. Such complex behavioral processes
encompass interactions among many neuronal systems involving
other neurotransmitters as well as peptides. Clearly, 5-HT systems
are contributory to a variety of important behavioral functions. In
linking serotonin and the serotonergic neural network to behav-
ior, it is necessary to recognize that behavior is sensory–motor
integration and therefore, the serotonergic system is anatomically
organized to play a major integrative role.
2.2. Serotonin and motor function
The most critical source of information about the connection
between activity in serotonergic neurons and motor function has
been provided by studies of drug induced serotonergic hyper-
and hypo-activity. A number of investigations have shown that
pharmacological over-stimulation of serotonergic system can gen-
erate a hyper-motility syndrome [19] and, in addition, reverse
neuroleptic-induced hypo-motility [20]. In that excessive seroto-
nergic activity generates a behavioral state of over-activation, the
expectation would be that inactivation of serotonin would be linked
to behavioral inactivation. Key support for serotonergic inactiv-
ity being linked to motoric inactivation has been provided by an
unusual source. Studies using unit activity recordings [21–23] in the
serotonergic neurons during the sleep–wakefulness cycle revealed
a positive correlation between serotonergic activity and wake-
fulness. A surprising and startling observation provided by this
research was the finding that serotonergic unit activity became
essentially silent during rapid eye movement sleep (REM). This
is of central importance with regard to the role of serotonin in
motor function in that during REM sleep, when there is intense
brain activity there is a movement paralysis [24,25]. This natural-
istic observational research uncovered a vital function of serotonin
in permitting REM sleep without the hazard of commensurate
motoric activation. In addition, these studies and other find-
ings indicate that serotonergic activity appears to be critical for
movement [26,27].
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Although 5-HT activity may be necessary for movement, other
sources of evidence have shown that serotonergic activity, by itself,
is insufficient to activate lower motor neuron activity [28,29]. This
necessary but not sufficient role for a neurotransmitter is certainly
not unique to serotonin, but rather has pointed-up the critical
contribution of serotonin as an enabler of movement through inter-
action with other neurotransmitter systems [30,31]. A number of
investigations have identified the importance of the interaction
between the excitatory amino acid (glutamate) and serotonin in
the modulation of lower motor neuron activity [32–34]. Impor-
tantly, this serotonin–glutamate linkage points to new directions
in the understanding and possible treatment of the devastat-
ing motor neuron disease of amyotrophic lateral sclerosis (ALS)
[35–37].
behavior and therefore serve as unconditioned stimuli that elicit
unconditioned drug responses. In preclinical Pavlovian psychos-
timulant drug conditioning studies, the simplest and most common
arrangement is to administer the drug prior to placement in a test
environment that serves as the contextual CS. This contrasts with
conventional Pavlovian conditioning in which the CS is restricted to
one sensory system (visual, auditory, tactile) and precisely paired
temporally to the UCS. In this discrete CS conditioning arrangement
the general testing environment provides the environmental con-
text for the conditioning and can contribute to the conditioning as
an occasion setter [52] rather than simply as a CS. In contrast, many
preclinical Pavlovian drug-conditioning studies have no explicit
discrete CS and the experimental test environment in which the
drug response occurs serves as the CS.

3.2. Paired/unpaired treatment design as a necessary feature in

2.3. Serotonin and sensory processing
studies of Pavlovian drug conditioning
While the critical contribution of serotonin neurons to move-
In the conduct of a Pavlovian conditioning protocol, a basic
ment in the “final common motor neuron pathway” is well
requirement is the use of the paired/unpaired design. As applied
established, the contribution of serotonergic neuronal activity to
to drug/test environment conditioning, the drug treatment used
higher brain function and to more global activity processes labeled
as the unconditioned stimulus is paired to a specific test environ-
as behavior are, needless to say, less evident. A vast literature has
ment complex and the same drug treatment is also administered
been developed involving a variety of pharmacological, genetic
but unpaired to this cue complex and serves as a control group. The
knockouts and lesion manipulations. Some recent reports, how-
unpaired control group is needed to rule out non-specific response
ever, appear to suggest an important and new way to understand
sensitization effects induced by the drug treatment such that the
the critical contribution of the serotonergic system to behavior and
response effect labeled as a conditioned response is not simply a
conditioning. A key finding is in the reports of Müller et al., and Pum
generalized elevated responsiveness to many different non-drug
et al. [38–40].
stimuli. This is of particular importance with regard to psychos-
Analogous to the electrophysiolgical findings of Jacobs and
timulant drugs in that the unconditioned drug response is often
Fornal [41], these investigators employed an in vivo monitor-
manifested in behavior as a heightened level of spontaneous behav-
ing of serotonergic activity using microdialysis in sensory cortex
ior. Thus, the conditioned response is not a unique behavioral
to monitor serotonin release in the awake behaving rat. Criti-
response but rather an increase in spontaneous behavior as com-
cally, this in vivo recording technique as with electrophysiological
pared to a control group; and, this differential is the marker for a
unit recordings involved monitoring of 5-HT systems without
conditioned drug response. Thus, in psychostimulant drug condi-
disrupting normative ongoing 5-HT activity. These microdialysis
tioning the paired/unpaired design is required to make an inference
investigations showed that drugs (e.g., cocaine) that increase sero-
of conditioning.
tonin extracellularly in primary sensory areas of the neocortex such
as the visual cortex. Additionally, these investigations showed that
3.3. Critical differences of drug conditioning from simplified
a behaviorally significant visual stimulus (i.e., flashing light) also
standard models of Pavlovian conditioning
increased serotonin release in the visual cortex and this release
was accompanied by increased behavioral activation. Importantly,
Typically, drug conditioning protocols differ from the standard
these microdialysis findings have provided a new way to concep-
Pavlovian conditioning paradigm in that the drug treatment (the
tualize the role of serotonin in sensory/motor function by showing
UCS) precedes rather than follows the CS but critically the UCS and
that activation of the serotonergic system can directly modulate
the CS overlap extensively so that drug conditioning can be con-
sensory information at the level of the sensory areas of the neo-
sidered a kind of reverse Pavlovian delay conditioning. While the
cortex, and in this way modify the salience or significance of
drug UCS precedes the contextual CS this arrangement is unlike
environmental stimuli. Clearly, if the salience of the stimulus is
backward conditioning in that the onset of the psychostimulant
enhanced then the behavioral response will also be amplified. By
UCS drug effect is not followed by an inconsequential stimulus
having a modulating influence upon direct primary sensory stimuli
as in conventional conditioning but rather the psychostimulant
at the cortical level and on the response output systems, serotonin is
drug effect induces sensory/motor activation so that the envi-
situated to have a pivotal role in reactivity to external environmen-
ronmental context is transformed into a highly salient stimulus
tal stimuli by modulation of sensory information in sensory cortices
complex by the drug UCS. Another deviation of this drug condition-
as well as efferent motor neurons in the spinal cord; thereby, pro-
ing from conventional Pavlovian conditioning is that the UCS drug
moting action, emotion and memory storage.
treatment can last for a substantial duration and occurs in a test
environment serving as the CS. Consequently, the CS–UCS overlap is
3. Psychostimulant drug conditioning substantially longer than for the conventional Pavlovian discrete CS
conditioning protocol. Perhaps, the most straightforward drug con-
3.1. Psychostimulant drug conditioning and high abuse liability ditioning entails the use of psychostimulant drugs such as cocaine,
amphetamine, apomorphine etc. that evoke an unconditioned drug
Psychostimulant drugs have a well-documented high abuse lia- response of hyperactivity and induce a conditioned drug response
bility. In contrast to many drugs that induce tolerance effects, the of hyperactivity relative to the unpaired control group. While this
repeated use of psychostimulants induce sensitization and con- drug conditioning paradigm can have complications that are not
ditioned drug effects that enhance the drug effects [42–51]. In completely resolved with the paired/unpaired experimental design
terms of Pavlovian conditioning, psychostimulant drugs are admin- [53,54] there is little doubt that this is a reliable drug condition-
istered to animals by an experimenter independent of the animal’s ing effect [55]. The use of a locomotor stimulant drug response to

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investigate conditioning has the advantage that the conditioned
response fulfills the basic element in Pavlovian conditioning in
that conditioned response is similar to the unconditioned response
albeit as an attenuated version [55].
3.4. Paradigms and preparations in the study of Pavlovian drug
conditioning
Perhaps the most common use of Pavlovian conditioning to
examine drug effects has been the use of the Conditioned Place Pref-
erence (CPP) protocol. While the CPP conditioning procedure is the
same as that used in the conditioning of the locomotor stimulant
drug response a major difference in the case of CPP is that there
is not a directly observed drug response but rather a hypothesized
positive hedonic drug response. This drug response is then subse-
quently validated or invalidated in a CPP test. Thus, if the animal
displays a preference for the environment in which the drug treat-
ment was administered it is then inferred that the drug treatment
did indeed induce a positive hedonic state and that this effect was
conditioned to the associated contextual cues. While the CPP effects
are of significance for the development of an improved understand-
ing of behavioral mechanisms that can initiate drug addiction this is
not a useful model for the study of drug conditioning in that the UCR
is not directly measurable but rather is inferred from the occurrence
of the CR (i.e., the CPP). The psychostimulant induced locomo-
tor activation response conditioning model has the advantage for
the study of drug conditioning in that the UCR and the CR are
directly observable and can be quantified in terms of distance mea-
sures. Indeed, there have been many reports of the conditioning of
the locomotor stimulant response of a number of psychostimulant
drugs particularly drugs that have dopamine agonist effects such
as cocaine, amphetamine and apomorphine [56].
3.5. Characteristic features of the Pavlovian drug conditioning
hyper/hypo-activity model
In this conditioning paradigm, the environment in which the
drug response occurs becomes the conditioned stimulus and the
drug-like response subsequently elicited by the environment in
a non-drug test is the conditioned response. Indeed, drug con-
ditioning has become a major source of Pavlovian conditioning
experimentation [56]. Alternatively, if drug administration is made
contingent upon a response emitted by the subject, then, the drug
is used to promote instrumental conditioning.
4. Methodological issues in the use of experimental
manipulations of serotonin to study psychostimulant drug
conditioning
4.1. Introduction
In that Pavlovian conditioning entails the formation of a new
linkage between sensory and motor responses, serotonergic sys-
tems would appear to have a critical role in this basic conditioning
process. Any attempt to assess the contribution of a neurotrans-
mitter system such as the serotonergic system involving as many
as 14 different types of receptors in terms of how the conditioning
processes may be affected is a formidable challenge. In addition,
there are the complexities of ruling out experimental manipula-
tions of the serotonergic system that can produce alterations in
sensory/motor processes with major indirect effects upon condi-
tioning. Such sensory/motor effects are not on the associative or
learning dimension but rather on the UCS, CS and/or the UCR, CR.
Thus, the attribution of associational effects to the experimental
manipulation of the serotonin system is indeed a daunting chal-
lenge. In this regard it is quite evident that in typical instrumental
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5
conditioning it is particularly difficult to unravel possible treatment
effects upon sensory/motor processes in that the acquisition of the
learned behavior relies upon the spontaneous behavior of the ani-
mal and is not in control of the experimenter. In psychostimulant
drug conditioning of psychomotor effects, however, the CS and the
UCS are under control of the experimenter and possible effects of
a serotonin treatment upon the CS and UCS can be determined.
In that the test environment serves as the CS and the degree to
which such a treatment may impact upon reactivity to the CS can
be assessed by the measurement of locomotor activity level in the
test environment. The serotonin effect upon the UCS can also be
assessed by the measurement of the motoric stimulation induced
by the psychostimulant drug. Only if the CS and UCS are unaffected
by the serotonin treatment can the possible serotonin effects upon
the CS–UCS association be investigated.
4.2. Methods for the behavioral analysis of serotonin
manipulations: “knockout” preparations
A basic approach to a functional analysis of neurotransmitter
systems is to perform experimental manipulations that selectively
impair neuronal activity in one neurotransmitter system and to
subsequently assess the impact of the treatment upon behavior. We
will briefly examine basic issues that need to be considered first in
linking experimental alterations in 5-HT to behavior. For example,
the recent utilization of “knockout” mouse preparations has pro-
vided one approach. While an animal model lacking 5-HT neurons
is non-viable, it is possible to create viable knockouts lacking in par-
ticular 5-HT receptor subtypes (e.g., the 5-H1B receptor) [57,58].
Although behavioral changes can be observed in these prepara-
tions, the impact cannot be easily ascribed simply to the of loss
of a specific receptor subtype in that many compensatory changes
in 5-HT neurons and other transmitter systems occur which make
it difficult to connect any of the observed behavioral changes to the
absence of the receptor subtype versus the compensatory changes
to this missing receptor subtype. Furthermore, this type of genetic
subtraction manipulation affects the expression of the 5-HT recep-
tors throughout the brain. Recent developments in selective gene
editing may obviate much of the above criticism; however, this
new technological breakthrough does not have sufficient experi-
mental exploration to allow sufficient basis for evaluating this more
advanced tool in the manipulation of serotonin or other transmitter
receptors [59,60].
4.3. Methods for the behavioral analysis of serotonin
manipulations: “lesion” preparations
In order to assess 5-HT involvement in complex behavioral
effects, it is necessary to selectively manipulate 5-HT neurons that
project to the forebrain from the dorsal and median raphe nuclei.
Indeed, there have been a large number of studies conducted in
which dorsal and median raphe nuclei have been damaged by
various lesion methods, including electrolytic- and neurotoxin-
induced lesions. In view of the relatively inaccessible location
and the elongated topography of the raphe nuclei, it is difficult
to produce virtually complete lesions of these nuclei using non-
specific lesion techniques such as electrolytic and radio-frequency
lesion procedures without also damaging surrounding reticular
formation and other neuronal tissue. Even more selective lesion
methods involving intracerebral injections of neurotoxins such as
5,7-dihydroxytryptamine infusions (5,7-DHT) can produce asym-
metric and incomplete lesions of 5-HT neurons, and can also
produce non-5-HT neuronal damage.
In other neurotransmitter systems in which there is extensive
axonal collateralization (e.g., dopamine), it is well documented that
neuronal cell loss in the rat must be severe; i.e., >90 percent [61]
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to produce persistent behavioral deficits. Additionally, it is long
known that in the case of human degenerative diseases such as
Parkinson disease (dopamine neurons) and amyotrophic lateral
sclerosis (ALS) (ventral horn motor neurons) the observable behav-
ior deficits tend to occur when cell losses are in the order of 60–80
percent [62,63]. The difficult requirement to produce severe cell
loss restricted selectively to dorsal and median raphe nuclei has
diminished the utility of lesion methods. Also, there is no human
neurodegenerative disorder syndrome that is specifically linked to
the dorsal and median raphe serotonin neurons in the expression
of the neurological deficit profile.
4.4. Behavioral assessment methods: incentive/reward
preparations
Coupled to the intrinsic experimental difficulties in the use of
an animal model preparation, in which the dorsal and median
raphe nuclei are selectively destroyed as well as in animal mod-
els using gene “knockouts” is the related problem of the necessary
and appropriate behavioral methods to detect and measure the
effects of such manipulations. In rodent models of learning and
memory, it is critical to differentiate between learning and per-
formance variables. For example, in a reward-based learning task,
an experimental manipulation that alters the effectiveness of the
reward used in the task would clearly have a performance effect in
that the capacity to acquire the behavioral response may be intact
but the incentive to perform would be changed.
4.5. Behavioral assessment methods: aversive learning
preparations
In addition to appetitive reward paradigms, there are also
widely used aversive learning paradigms in which painful stimuli
such as electric foot-shock are administered to motivate learning.
The most commonly used tasks in animal studies are passive avoid-
ance tasks in which the behavior learned is response inhibition
of spontaneous behavior that leads to footshock, and, conversely,
active avoidance tasks in which the learned behavior is the acqui-
sition of an anticipatory escape response to avoid a foot-shock.
Clearly, experimental manipulations of 5-HT neurons that alter
sensitivity to the aversive stimulus and/or response output impact
upon such learning tests [64]. A 5-HT treatment that reduces
response output would facilitate passive avoidance learning but
impair active avoidance; and conversely a 5-HT manipulation that
increases response output would have the opposite effect. Thus, a
modification in reactivity to the aversive stimulus used to moti-
vate behavior could enhance/impair learning in both types of
tasks.
4.6. Behavioral assessments of transmitter manipulations
The effects of transmitter manipulations upon the motivational
properties of a reward/aversive stimulus used in a learning task
might be performance effects rather than direct effects upon asso-
ciative processes. Needless to say, profound effects upon response
output are obvious; however, even if, for example, a seroto-
nergic treatment leaves an animal without an apparent motor
impairment, there could be subtle changes such as the tendency,
rather than the capacity, to respond that are more difficult to
detect.
What is needed is a simple measure of the spontaneous base-
line activity level of an animal. In fact, a wide variety of behavioral
measurement methods have been developed in an effort to make
the basic assessment of baseline response output. One of the most
widely used methods in rodents is to place a test animal in an open
arena and record movement during a fixed time interval. While
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conditioning. Behav Brain Res (2014), http://dx.doi.org/10.1016/j.bbr.2014.10.038
automated systems incorporating photo beam interruptions have
been widely used, this procedure provides only an indirect measure
and does not capture the full range of behavioral movement (e.g.,
rearing, grooming, locomotion). Direct observations in scoring of
responses combined with videotape scoring, while labor intensive,
can provide a reliable and comprehensive capture of behavioral
activity. Newer automated digitized video image recordings of dif-
ferent behavioral responses increasingly are becoming available
to avoid the complications of reliability in observer evaluations of
behavior.
4.7. Significant test environment variables
Detailed recording of behavior is necessary but not sufficient
in that many test environment variables (boundary conditions)
need to be considered/identified and held constant (e.g., arena
size, lighting, and test duration and frequency). Lighting condi-
tions are important factors in that white light to rodents is an
aversive stimulus so that a 5-HT manipulation could impact upon
activity indirectly by diminishing or enhancing the aversive qual-
ity of the white light. These concerns can be avoided by the use
of red light in that rodents are relatively insensitive to these
longer wavelengths. While the diurnal cycle is a substantial vari-
able affecting rodent activity, for acute testing of activity level, dark
or red light conditions favor less emotional stress to interfere with
behavioral activation, and therefore may provide less problematic
testing conditions for rodents [65]. Besides behavioral record-
ing methods and lighting, it is necessary to consider the testing
protocol.
4.7.1. Novelty/familiarity variables
A major determinant of behavioral activity in a test arena is the
novelty/familiarity of the test environment [66]. The more novel
the environment, the greater the behavioral activity expressed in
the control treatment groups. Typically, treatment manipulations
designed to modify brain serotonin do not use a novel environment
to assess serotonergic effects upon response output. A considera-
tion relevant to the use of a novel environment is that the control
group activity level is high, so it becomes more difficult to detect
an increase in activity. In several studies, however, with stimu-
lant drugs (i.e., cocaine), we have shown that cocaine at a modest
dose level does not induce behavioral abnormalities or increases
in movement stereotypes in a novel environment [53]. This result
indicates that initial activity in a novel environment does not create
a response output ceiling effect, and could be an effective technique
to assess 5-HT manipulations on behavioral activation capacity. On
the other hand, changes in overall motility can be misleading in
that activity changes can occur within a test session (i.e., habitu-
ation) and 5-HT treatments potentially can modulate locomotor
activation by affecting the capacity to habituate to an environ-
ment.
In intact control animals, locomotor activation is highest in the
initial phase of testing in a novel environment but then progres-
sively declines as the animal acquires familiarity with the test
environment. A 5-HT manipulation, therefore, could increase loco-
motor activation by reducing the capacity to habituate to the test
environment cues. Consequently, an overall increase in activity
level by itself is not a sufficient assessment of response output
effects of treatment variables since it may indicate an inability to
acquire information rather than an overactive response system. In
order to avoid this possibility, another approach is to place the
animal in a test environment for a substantial time period prior
to performing a manipulation upon the 5-HT system. In that the
pre-exposure lowers the response output of the control treatment
to a stable level of response output, this protocol can be seen
as an effective way to detect a change in response output. An
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increase in behavioral activity in such a testing protocol, however,
does not necessarily imply direct activation of response systems
in that the increase could also be attributed to an effect of the
5-HT manipulation upon the acquired habituation/familiarity to
the test environment so that the environment now reverts back
to being more novel and in this way evokes an increase in response
output. Thus, the increase in response output could be a direct
motoric effect or it could be indirect by virtue of an impact on stim-
ulus processing or on memory of context familiarity/habituation.
Altogether, these considerations make it evident that even the
seemingly simple measurement of response output can be a com-
plex task.
4.7.2. Stimulus processing
In a study [67] pertinent to the issue of the role of serotonin in
processing external stimuli, we manipulated serotonin availabil-
ity by the use of low auto-receptor preferring dose levels (0.01,
0.025, 0.05 mg/kg) of the selective 5-HT1A agonist (8-OHDPAT)
that decreases brain 5HT metabolism [68] and 5-HT1A antago-
nist (WAY-100635) which can enhance ongoing serotonin activity
[69] in a testing paradigm with rats in which we measured the
behavioral responsiveness to a small novel object placed at the
center of an open field. A novel object is a highly salient stimu-
lus that is responded to spontaneously thereby circumventing the
complications associated with stimulus salience acquired through
extensive reward training. Importantly, we found that the 5-HT1A
agonist 8-OHDPAT, at an autoreceptor dose level (0.025 mg/kg),
diminished the animal’s behavioral responses to the novel object
stimulus without impairing spontaneous motor activity; in con-
trast, the selective auto-receptor antagonist, WAY-100635, did not
alter spontaneous locomotor activity, but increased responsive-
ness to the novel object at the 0.025 mg/kg dose level. As the
dose levels of these drugs were increased to 0.05 mg/kg the effect
became exaggerated; and, in the case of the agonist 8-OHDPAT,
it virtually eliminated responsiveness to the novel object as well
as partially decreasing motor activity. It is readily apparent from
these findings that such global alterations in serotonergic activ-
ity produce profound changes in sensory/motor behavior so that
it is not possible to determine the contribution of serotonin to
associational mechanisms using conventional conditioning proto-
cols.
4.8. Drug stimulus effects on conditioning
While there have been a number of reports that suggest that var-
ious 5-HT receptor subtypes can modify the reward value of drugs
such as cocaine [70,71], we have shown in several reports that the
serotonergic drug effects may be the result of internal drug stimu-
lus effects. Like many psychoactive drugs [72–74], centrally active
serotonergic drugs are presumed to have discriminative stimulus
properties that are not necessarily evident in the motor behavior
particularly when given in combination with cocaine wherein the
cocaine treatment drives motor output. In fact, we have shown
[72,73], that cocaine context dependent sensitization and con-
ditioned effects can be gated by the drug stimuli generated by
serotonergic drugs such as the selective 5-HT1A agonist 8-OHDPAT.
In these studies we employed a typical protocol for assessing
drug treatment effects upon cocaine sensitization and conditioning
wherein the control group receives saline plus cocaine treatments
and the experimental group receives a drug treatment (e.g., 8-
OHDPAT) in combination with cocaine at a dose level that does not
affect the locomotor stimulant response induced by cocaine. Sub-
sequently, a challenge test is performed in which the 8-OHDPAT
plus cocaine group and the saline plus cocaine group are given a
saline plus cocaine test. In line with the a role for the serotonergic
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conditioning. Behav Brain Res (2014), http://dx.doi.org/10.1016/j.bbr.2014.10.038
7
system in the acquisition of cocaine sensitization only the saline
plus cocaine control group exhibited a sensitization effects. In order
to assess the possible contribution of 8-OHDPAT to the stimulus
complex we next, gave both groups a second challenge test in which
both groups were given 8-OHDPAT plus cocaine. In this challenge
test the experimental group (8-OHDPAT plus cocaine) exhibited a
sensitization effect but the saline plus cocaine control group now
tested with 8-OHDPAT plus cocaine did not exhibit a sensitization
effect. This experiment demonstrates the way in which the stim-
ulus properties of a drug treatment can become incorporated into
the contextual stimulus complex and acquire conditioned stimulus
properties. Consequently, conducting a sensitization challenge test
that only includes a saline sensitization test creates the possibil-
ity that a false positive result can be obtained and can lead to the
inappropriate conclusion that the neurotransmitter system modi-
fied by the serotonin drug treatment in this example prevents the
acquisition of cocaine sensitization. Thus, it is critical to incorpo-
rate an assessment of possible drug stimulus effects to determine
whether or not a drug manipulation of the serotonin system that
attenuates or blocks psychostimulant conditioning has altered the
association process or has contributed to the conditioned stimulus.
Indeed there are a number of reports using 5-HT2C agonist drugs
[70,71] that suggest that the activation of these receptor subtypes
appear to be able to attenuate the reward properties of cocaine
as well as CS-induced drug seeking behavior. Whether or not these
effects are also explicable by the introduction of drug stimulus cues
remains to be determined.
4.9. Drug stimulus effects in testing for conditioned drug
treatment effects
In a number of our studies on cocaine conditioning and sen-
sitization, we demonstrated that the decrements in conditioning
and sensitization linked to the serotonergic drug treatments were
attributable to the drug stimulus properties of these drugs in that
the conditioning and sensitization decrements could be restored if
the drug stimuli were provided [72,73]. For example, we showed
that the partial 5-HT1A agonist, Buspirone, by itself, had no effect
upon spontaneous behavior, nonetheless, could eliminate a cocaine
conditioned locomotor response either (a) when given prior to a
conditioning test following the induction of cocaine conditioning
or, (b) if buspirone were given in combination with cocaine during
the induction of conditioning, but buspirone were not given before
the subsequent test for conditioning. The critical experimental step
we took was to then administer Buspirone to the Buspirone plus
cocaine group that did not exhibit a cocaine conditioned response
in a standard saline conditioning test. In response to the Buspirone
pre-treatment, this group then expressed a conditioned response
that was equivalent to the conditioned response of the group that
had received cocaine without Buspirone during the conditioning
induction phase.
Altogether, these results are readily explicable in terms of the
stimulus properties of the drug such that the cues generated by
drug stimuli can be a critical element in the conditioned stimu-
lus complex. If not present during conditioning, the addition of the
drug cues prior to a conditioning test can alter the CS so that it is no
longer effective. If present during induction of conditioning, then
their presence or absence in the conditioning test determines the
occurrence or absence of the CR. In that virtually all drug condi-
tioning studies using serotonergic drugs have not employed this
critical testing procedure to assess the possible contribution of the
serotonergic drug stimuli to the conditioned stimulus it is difficult
to interpret the nature of the contribution serotonergic drugs to
psychostimulant drug conditioning.
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5. A new model to assess the role of serotonin in
conditioning
5.1. Conditioning and the memory trace
Recently [74,75], we re-conceptualized Pavlovian conditioning
from a CS–CR formulation to a CS–memory trace–CR (CS–MT–CR)
process framework formulation. Importantly, our current stud-
ies employing a design based on this re-conceptualized Pavlovian
model, point to a new perspective regarding the plasticity of an
established memory trace. Results of these studies suggest that
viewing the CS as activating a memory trace that is expressed as the
CR, can offer an alternative strategy to assess if a treatment impacts
the CS–CR linkage.
5.1.1. Memory trace and consolidation phase
For a long time it was thought that a brief dynamic phase in
the formation of a memory trace was thought to exist only at
the time of the formation of a new memory trace and that only
during this memory forming consolidation period could an inter-
vention prevent/enhance memory formation. This consolidation
aspect of memory has been long recognized [76,77] in a variety of
learning paradigms in which manipulations ranging from electro-
convulsive shock (ECS) [78] to protein synthesis inhibitors [79–81]
to glucose [82,83], have been administered during a short post-
trial temporal interval following a learning trial and have been
shown to block/enhance retention. These same treatments admin-
istered after a longer temporal interval (e.g., 2 h) have no effect upon
retention of the learning experience. Thus, a period of memory
vulnerability followed by invulnerability has led to a general con-
sensus that after a brief post-trial interval, memory traces become
consolidated and incorporated into the physical structure of the
brain.
5.1.2. Memory trace and reconsolidation phase
Current analyses of memory, however, have suggested that the
memory trace is not hard wired; but that memory traces each
time they are reactivated once again become labile and sensitive
to modification [78]. This alternative conceptualization creates the
possibility that an established memory can be re-activated by cue
re-exposure and then modified. It is labeled as reconsolidation. In
fact, recent studies have shown that amnestic post-trial treatments
that follow shortly after re-exposure of animals to cues associated
with cocaine can impair the drug memory traces [84].
5.2. Neurotransmitter inhibition and reconsolidation of
conditioning
In our most recent studies, we have shown that repeated
pairing of the D1/D2 agonist apomorphine, at dose levels that
elicit hyper-locomotion with placement in an open-field, gen-
erates both context specific cue conditioned hyper-locomotion
and context specific sensitized drug induced hyper-locomotion
effects [47,49,51,85]. In line with the extant literature, extinc-
tion eliminates the conditioned apomorphine hyper-locomotion
response but not the sensitized apomorphine hyper-locomotion
response. Surprisingly, we recently reported that immediate post-
trial administration of a low auto-receptor dose of apomorphine
following brief exposure to the conditioned stimuli in a memory
re-consolidation paradigm eliminated not only the conditioned
response but also the sensitized apomorphine hyper-locomotion
response [75,74]. The same post-trial treatments were without
effect if they were administered after a delay of 15 min by which
time conditioning would have been re-consolidated. In addition,
the same post-trial treatments given repeatedly to groups that were
not conditioned had no discernable effect upon behavior. Thus, the
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post-trial treatments were selective to the conditioned association
that was re-activated by a brief exposure to the conditioned stimuli.
In this way we were able to demonstrate that the CS–CR condi-
tioned drug bond could be substantially modified by a post-trial
drug treatment that selectively targeted a specific neurotransmitter
system.
5.3. The post-trial drug treatment model as a tool to assess
specific serotonin receptor subtypes implicated in
psychostimulant conditioning
The post-trial drug treatment protocol seems well-suited to
assess the possible contribution of the serotonergic system to drug
induced conditioned effects by having the drug treatment be deliv-
ered to coincide with the re-consolidation phase of the conditioned
memory trace. While the overall contribution of the serotonergic
system can be assessed with global 5-HT treatments such as auto-
receptor doses of 8-OHDPAT, it will be feasible to determine the
potential critical importance of specific receptor subtypes such as
the 5-HT2C receptor that have been implicated in psychostimu-
lant conditioning and in motivational incentive salience effects in
cue-elicited drug seeking behavior [70,71].
Critically, the use of the post-trial model eliminates possible
performance related drug effects from the interpretation of the
results in that the exposure to the CS trials are all conducted with-
out the serotonergic drug treatments and any possible post-trial
carry over drug effects are controlled for by the use of delayed post-
trial treatments that are administered after the re-consolidation
has occurred. Thus, the memory re-consolidation model described
above offers a potential protocol for future investigations by dif-
ferentiating serotonergic system involvement in the associational
dimension of the conditioning process from a potential impact on
performance by altering sensory/motor function.
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Please cite this article in press as: Carey RJ, Damianopoulos EN. Serotonin and conditioning: Focus on Pavlovian psychostimulant drug
conditioning. Behav Brain Res (2014), http://dx.doi.org/10.1016/j.bbr.2014.10.038