Enhancing Effects of Sound on Methamphetamine-Induced Behavioral Aberrations in the Rat: A Model of Relapse of Schizophrenia-Like Symptoms

Abstract: Conditioning of methamphetamine (MAP)-induced behavioral aberrations, hyperactivity (HA), stereotyped (SB) and bizarre behavior (BB) in rats were examined using Pavlovian conditioning methods. A speaker-sound (SS) was used as the conditioning stimulus (CS) and MAP-induced behavior as the unconditioned stimulus (UCS). The rats belonging to the Sound Group were repeatedly administered with MAP or saline paired with SS in an observation room and those in the Soundless Group with MAP only in the housing room.
After chronic administration with MAP, conditioned behavioral responses (CBR) such as HA, SB, and BB were observed more intensely in the Sound Group than in the Soundless Group while being subjected to SS. CBR were greatest on the 30th day after the last MAP administration and they did not disappear for 120 days.
CBR were considered as analogous to the spontaneous relapse of schizophrenia-like symptoms.
Furthermore, as a certain behavior which had been conditioned could easily be observed as CBR with both specific (SS) and nonspecific environmental cues (handling), it is difficult to distinguish the withdrawal behavioral effects of MAP that lacks those of CBR.     

Behaviorally conditioned immunosuppression in the rat is regulated via noradrenaline and beta-adrenoceptors

Using Cyclosporin A (CsA) as an unconditioned stimulus has previously demonstrated that behaviorally conditioned inhibition of splenocyte proliferation and cytokine production is mediated via the splenic nerve. Therefore, we currently examined the adrenergic modulation of conditioned suppression of splenocyte function. Chemical sympathectomy via 6-OHDA completely blocked the conditioned suppression of splenocyte proliferation to mitogens and cytokine (IL-2, IFN-gamma) production. Furthermore, administration of beta-adrenoceptor antagonist propranolol abrogated the conditioned effect on splenocyte proliferation. Supporting the position that conditioning is beta-adrenergic-dependent, addition of beta-adrenoceptor agonist, but not alpha-adrenoceptor agonists, to splenocytes in vitro mimicked the conditioned suppression of splenocyte functions, with these effects blocked by propranolol. Therefore, these data indicate that behavioral conditioning of splenocyte function in the rat is regulated by the sympathetic nervous system, predominantly via beta-adrenergic mechanisms.     

A quantitative method for assessing of the affective component of the pain: conditioned response associated with CO2 laser-induced nocifensive reaction

Sensory and affective components are included in the overall behavioral manifestation in a nocifensive reaction. We have developed a behavioral model using classical conditioning to differentiate the affective component from the sensory responses following a thermal noxious stimulus. In laser-pain conditioning, free moving rats were trained to associate a tone (conditioned stimulus, CS) and short CO2 laser pulsation (unconditioned stimulus, US). A monotonous tone (800 Hz, 0.6 s) was delivered through a loudspeaker as the CS. CO(2) laser pulses (5 W at 100 ms in duration) applied to the hind paw were adopted as the US. The CS-US interval was 0.5 s. The conditioned responses as quantitatively measured by their body movement were developed over a period of 40 CS-US pairings. These conditioned responses were found retained when the rats were tested by presenting CS alone, immediate to and 24 h subsequent to training. The conditioned responses however diminished significantly following both morphine and buspirone treatment. This method demonstrated that neutral auditory stimuli could form association with unlearned nocifensive responses evoked by noxious CO2 laser pulses stimuli. Thus, the assessment of conditioned response may be a valuable tool for the measurement of the affective component of nociception.     

Classical conditioning analog enhanced acetylcholine responses but reduced excitability of an identified neuron

Although classical and operant conditioning are operationally distinct, it is unclear whether these two forms of learning are mechanistically distinct or similar. Feeding behavior of Aplysia provides a useful model system for addressing this issue. Both classical and operant appetitive behavioral training enhance feeding, and neuronal correlates have been identified. Behavioral training was replicated by in vitro analogs that use isolated ganglia. Moreover, a single-cell analog of operant conditioning was developed using neuron B51, a cell important for the expression of the conditioned behavior. Here, a single-cell analog of classical conditioning was developed. Acetylcholine (ACh) mediated the conditioned stimulus (CS)-elicited excitation of B51 in ganglia and mimicked the CS in the single-cell analog of classical conditioning. Pairing ACh with dopamine, which mediates the unconditioned stimulus in ganglia, decreased the excitability of B51, and increased the CS-elicited excitation of B51, similar to results following both in vivo and in vitro classical training. Finally, a D1 dopamine receptor (D1R) agonist failed to support classical conditioning in the cellular analog, whereas D1R mediates reinforcement in operant conditioning.     

Conditioned place aversion with interleukin-1beta in mice is not associated with activation of the cytokine network

Several distinct findings argue in favor of conditioning of some components of the acute phase reaction. However, the possibility of a conditioned cytokine response has not been assessed. In the present study, this possibility was tested by submitting mice to place aversion conditioning with interleukin-1beta (2 microgram/mouse, ip) as the unconditioned stimulus and an odorous compartment of a two-compartment cage as the conditioned stimulus. After two pairings, conditioned mice developed place aversion towards the odorous compartment. However, this behavioral conditioning was not accompanied by any alteration in peripheral (spleen) and brain (hypothalamus) cytokine levels (interleukin-1, interleukin-6, and interleukin-10). These data do not support the possibility of conditioned alterations in the cytokine network.     

Classical conditioning in rabbits using pontine nucleus stimulation as a conditioned stimulus and inferior olive stimulation as an unconditioned stimulus

Classical conditioning of skeletal muscle responses was accomplished by pairing microstimulation of the pontine nuclei as a conditioned stimulus (CS) with microstimulation of the dorsal accessory olive as an unconditioned stimulus (US). A conditioned response identical in form to the behavioral response elicited by the olivary stimulation was established when the CS was forward paired with the US, and behavioral extinction occurred with CS-alone presentations or unpaired CS-US presentations. Conditioned responding could not be established or maintained when the CS and US were simultaneously presented or when the US preceded the CS (i.e., backward paired). Complete lesions of the interpositus nucleus abolished both conditioned and unconditioned responses. These findings support the idea that plasticity associated with classical conditioning of skeletal muscle responses occurs in regions of the cerebellum that receive convergent CS and US input.     

Simple and Complex Spike Firing Patterns in Purkinje Cells During Classical Conditioning

Classical blink conditioning is known to depend critically on the cerebellum and the relevant circuitry is gradually being unravelled. Several lines of evidence support the theory that the conditioned stimulus is transmitted by mossy fibers to the cerebellar cortex whereas the unconditioned stimulus is transmitted by climbing fibers. This view has been dramatically confirmed by recent Purkinje cell recordings during training with a classical conditioning paradigm. We have tracked the activity of single Purkinje cells with microelectrodes for several hours in decerebrate ferrets during learning, extinction, and relearning. Paired peripheral forelimb and periocular stimulation, as well as paired direct stimulation of cerebellar afferent pathways (mossy and climbing fibers) causes acquisition of a pause response in Purkinje cell simple spike firing. This conditioned Purkinje cell response has temporal properties that match those of the behavioral response. Its latency varies with the interstimulus interval and it responds to manipulations of the conditioned stimulus in the same way that the blink does. Complex spike firing largely mirrors the simple spike behavior. We have previously suggested that cerebellar learning is subject to a negative feedback control via the inhibitory nucleo-olivary pathway. As the Purkinje cell learns to respond to the conditioned stimulus with a suppression of simple spikes, disinhibition of anterior interpositus neurons would be expected to cause inhibition of the inferior olive. Observations of complex spike firing in the Purkinje cells during conditioning and extinction confirm this prediction. Before training, complex spikes are unaffected or facilitated by the conditioned stimulus, but as the simple spike pause response develops, spontaneous and stimulus-evoked complex spikes are also strongly suppressed by the conditioned stimulus. After extinction of the simple spike pause response, the complex spikes reappear.     

Contribution of the anterior cingulate cortex to laser-pain conditioning in rats

The emotional component of nociception is seldom distinguished from pain behavioral testing. The aim of the present study was to develop a behavioral test that indicates the emotional pain responses using the classical conditioning paradigm. The role of the anterior cingulate cortex (ACC) in the process of this pain conditioning response was also evaluated. In laser-pain conditioning, free moving rats were trained to associate a tone (conditioned stimulus, CS) and short CO₂ laser pulsation (unconditioned stimulus, US). Monotonous tone (800 Hz, 0.6 s) was delivered through a loud-speaker as CS. CO₂ laser pulses (5 W at 50 or 100 ms in duration) applied to the hind paw was adopted as US. The CS–US interval was 0.5 s. Laser-pain conditioning was developed during 40 CS–US pairings. CS and US pairing with 100-ms laser pulse stimuli was more effective in establishing conditioning responses than that of 50-ms stimuli. The conditioning responses remained, tested by presenting CS alone, immediate to and 24 h subsequent to training. The performance of laser-pain conditioning was significantly reduced after bilateral lesioning of the ACC. Similar results were also obtained by bilateral lesions of the amygdala. The conditioning responses were also diminished following morphine treatment. The association between a neutral stimulus and a noxious stimulus could be demonstrated in a Pavlovian conditioning test in free moving rats. Thus, the conditioned response may be employed as a measure of the emotional component of the nociception. It is also suggested that the ACC may play an important role in mediating this conditioning effect.     

Cortical evoked potential changes during classical conditioning of morphine dependence in rats

A classical conditioning experiment was designed to determine if a conditioned neural response would develop and persist in cortical evoked potentials elicited by a foreleg stimulus (CS+) that was paired with morphine administration during the development of dependence and subsequent withdrawal. A stimulus to the other foreleg (CS-) was presented explicitly unpaired with morphine delivery. After dependence was established, the rats were taken from the experimental chamber and withdrawn from morphine for 6 days in their home cages. Finally, during the testing phase, the animals were returned to the experimental chamber and the foreleg stimuli were presented. The CS+ was paired with either morphine or saline injections. Changes due to both morphine effects only and conditioning were observed. The conditioned response, however, was present only in the cortical evoked potentials recorded from those animals receiving contralateral foreleg stimulation as the CS+. The conditioned neural response persisted after withdrawal and was present in both the drug-free and morphine-intoxicated animals. These results provide support for the relapse theory that a nonextinguished conditioned response is retained after withdrawal. However, further experiments are necessary to determine if these conditioned responses can elicit drug-seeking behavior.     

Classical conditioning of endocrine effects

PURPOSE OF REVIEW: Classical conditioning is a form of associative learning, based on the pioneering work of I. P. Pavlov: due to its association with an unconditioned stimulus that induces an unconditioned response, an initially neutral stimulus will become a conditioned stimulus evoking a conditioned response in the absence of the unconditioned stimulus. One important area for the application of conditioning principles is the regulation of physiological systems in general, and endocrine responses and its concomitant changes specifically. Conditioned endocrine responses were predominantly addressed in animal studies so far, mainly examining conditioned insulin production (and blood-glucose change) and corticosterone release. RECENT FINDINGS: There are very few studies on classical conditioning of endocrine responses in the annual period of this review. The advancement, however, is that some are conducted with humans. Recently, as a new avenue, hormones and neurotransmitters have been examined as mediators of basic conditioning processes. Moreover, there is an increasing interest in gender-specific conditioning responses which are influenced by gonadal hormones. SUMMARY: Research on classical conditioning demonstrates that endocrine systems are adaptable to environmental signals. Likewise, the endocrine status of an organism (at least with regard to glucocorticoids and gonadal hormones) was shown to modify classically conditioned responses. Partly due to the high expenditure of conducting conditioning experiments, the quantity of studies is limited, but there is a need to extend this research to humans. In sum, the application of conditioning paradigms constitutes an important research tool for behavioral medicine as well as psychiatry to examine brain-behavior relationships.     

Conditioned responses in the visual cortex of dogs. II. During sleep

Repetitive electrical stimulation of the optic chiasm was used as a cue stimulus for alimentary conditioned lever-press in dogs. During sleep after conditioning, the cue was presented to test whether the conditioned response (CR) can be emitted during sleep and whether CR in visual cortex (VC) is due to change in excitability in VC. Though level-press behavior never occurred without accompanying EEG arousal responses, rate of occurrence periods of EMG (percent EMG) and rate of EMG in the lever-press forelimb increased in the conditioned animals. Amplitudes of components 4 and 5 of the evoked potential in VC enhanced during both slow-wave (SS) and fast-wave sleep (FS) with similar time course as in wakefulness. In addition, correlation between latency of the EMG and of maximum enhancement of the amplitudes which was obtained during wakefulness remained during these sleep stages. Enhancement of the amplitudes was greater during sleep than during wakefulness; no significant difference was found between SS and FS. These results support the hypothesis that the CR in VC is attributed to excitability change and suggest that CR, in the peripheral and central level, can be emitted during SS and FS periods in dogs.     

Different projections of the central amygdaloid nucleus mediate autonomic and behavioral correlates of conditioned fear

The purpose of the present study was to determine whether lesions of areas projected to by the central amygdaloid nucleus (ACE) would disrupt the classical conditioning of autonomic and/or behavioral emotional responses. The areas studied included 3 projection targets of the ACE: the lateral hypothalamic area (LH), midbrain central gray (CG) region, and bed nucleus of the stria terminalis (BNST). Lesions were made either electrolytically or by microinjection of ibotenic acid, which destroys local neurons without interrupting fibers of passage. Two weeks later, the animals were classically conditioned by pairing an acoustic stimulus with footshock. The next day, conditioned changes in autonomic activity (increases in arterial pressure) and emotional behavior ("freezing," or the arrest of somatomotor activity) evoked by the acoustic conditioned stimulus (CS) were measured during extinction trials. Electrolytic and ibotenic acid lesions of the LH interfered with the conditioned arterial pressure response, but did not affect conditioned freezing. Electrolytic lesions of the rostral CG disrupted conditioned freezing but not conditioned changes in arterial pressure. Ibotenic acid injected into the rostral CG reduced neither the arterial pressure nor the freezing response. Injection of ibotenic acid in the caudal CG, like electrolytic lesions of the rostral CG, disrupted the freezing, but not the arterial pressure response. Injection of ibotenic acid into the BNST had no effect on either response. These data demonstrate that neurons in the LH are involved in the autonomic, but not the behavioral, conditioned response pathway, whereas neurons in the caudal CG are involved in the behavioral, but not the autonomic, pathway. Different efferent projections of the central amygdala thus appear to mediate the behavioral and autonomic concomitants of conditioned fear.     

Modification of neuronal discharge along the ascending tectofugal pathway during visual conditioning

Visually conditioned heart-rate change in the pigeon has been developed as vertebrate model system for analysis of associative learning. The visual pathways transmitting the conditioned stimulus information were identified, and neurophysiological analyses during conditioning were then undertaken to determine if these pathways behave merely as input lines or undergo training-induced modification. After finding that the retinal output is invariant with training, we investigated the central visual pathways, beginning with the tectofugal pathway. During conditioning single neurons in the nucleus rotundus and ectostriatum, the thalamic and telencephalic relays of the tectofugal pathway, showed enhancement of their phasic light-evoked responses. In contrast, the initial phasic responses attenuated during non-associative training. The rate at which these discharge modifications developed paralleled the development of the behavioral response. Thus, the tectofugal pathway shows plasticity during conditioning and does not behave merely as an input channel for the conditioned stimulus.     

Effects of cannabidiol and diazepam on behavioral and cardiovascular responses induced by contextual conditioned fear in rats

Cannabidiol (CBD) is a non-psychotomimetic compound from Cannabis sativa that induces anxiolytic-like effects similar to diazepam in animal models of innate aversive behavior. However, the effects of CBD contextual conditioned fear have not been studied. Therefore, the aim of this work was to compare the behavioral and cardiovascular effects of CBD and diazepam, a prototype anxiolytic, in animals submitted to a contextual conditioned fear paradigm. Male Wistar rats were submitted to a 10min conditioning session (six footshocks, 2.5 mA, 3s, delivered at pseudo-random intervals). The behavioral and cardiovascular responses to the context were measured 24h later in a 10 min test session. Diazepam (2.5 mg/kg), FG-7142 (8 mg/kg), a benzodiazepine inverse agonist, or CBD (10 mg/kg) were administered i.p. before the test session. Conditioned rats submitted to the aversive context exhibited more freezing behavior and a larger increase in blood pressure and heart rate as compared to non-conditioned animals. These effects were attenuated by CBD and diazepam in the conditioned animals. These drugs did not have any effect in non-conditioned rats. FG-7142 treatment failed to change the behavioral and cardiovascular responses to the aversive context. In conclusion, the results suggest that CBD has anxiolytic-like properties similar to those of diazepam in a rat model of conditioned fear to context.     

Hippocampal plasticity during jaw movement conditioning in the rabbit

Hippocampal CA1 unit responses were recorded during classical conditioning of rhytmic jaw movements in New Zealand White rabbits. Training was accomplished using a 1 kHz tone as the conditioned stimulus (CS) and 1 ml of sweetened water as the unconditioned stimulus (US). The interstimulus interval was 250 ms. Daily sessions consisted of 48 paired trials and six tonealone test trials, with an intertial interval averaging 60 s. Controls were given explicity unpaired stimuli. Unit and behavioral conditioned responses developed very rapidly in the trained group, but did not occur in controls. Averaged unit poststimulus histograms showed a correspondence between rhytmic cell discharges and the periodicity of the behavioral conditioned response after training. The results are discussed in relation to a hippocampal role in the modulation of learned movement patterns.     

The effect of rewarding hypothalamic stimulation on behavioral and neural hippocampal responses during trace eyeblink conditioning in rabbit (Oryctolagus cuniculus)

Rabbits were trace-conditioned with a tone as a conditioned stimulus and an airpuff as an unconditioned stimulus. Electrical stimulation to the medial forebrain bundle in the lateral hypothalamus was delivered either before or after the tone-airpuff pair. The purpose of the present study was to test whether the effect of post-trial hypothalamic stimulation differed from the effect of pre-trial hypothalamic stimulation on trace conditioning in the same subjects. Additionally, hippocampal responses were measured during sessions to see if hypothalamic stimulation activated dopaminergic fibres and affected hippocampal cell functioning and thus learning. The results showed that behavioral nictitating membrane conditioned responses were acquired quickly and hippocampal multiple unit activity increased with post-trial hypothalamic stimulation in comparison to almost non-existent conditioned responses and activity changes in the pre-trial hypothalamic stimulation sessions. The results suggest that hypothalamic stimulation affects trace conditioning differently depending on its time of delivery during conditioning.     

Effects of lesions of cerebellar nuclei on conditioned behavioral and hippocampal neuronal responses

Rabbits were overtrained using classical conditioning of the rabbit nictitating membrane (NM)/eyelid response. Unilateral electrolytic lesions were then made through electrodes previously implanted in dentate and interpositus cerebellar nuclei ipsilateral to the trained (left) eye. Lesions caused a complete or near-complete abolition of conditioned behavioral responses on the ipsilateral side, but had no effect on unconditioned responses to corneal airpuff. When training was switched to the contralateral (right) side, animals learned within the first few trials, but did not relearn when training was returned to the left (lesioned) side. Control animals in which lesions spared the deep nuclei showed no such learning deficits.Lesions of cerebellar nuclei also abolished conditioned increases in hippocampal CA1 neural activity evoked by the tone conditioned stimulus in this paradigm. As with the behavior, training on the right (non-lesioned) side reinstated the conditioned neuronal response within the first few trials of training, even though behavioral responding on the lesioned side showed little or no improvement.These results indicate that the cerebellum is an essential structure for the behavioral expression of learning, and plays an important role in the generation of conditioned hippocampal responses observed in this paradigm.     

Pavlovian conditioning of endotoxin-tolerance in rats

The most fascinating example of the bi-directional interaction between the central nervous system (CNS) and immune system is the behavioral conditioning of immune functions. We therefore investigated the behavioral conditioning of lipopolysaccharide (LPS)-induced endotoxin tolerance using the taste aversion paradigm. The conditioned stimulus (CS) saccharin was paired with the unconditioned stimulus (UCS) LPS over a five (CONDl) or four (COND2) days learning trial. Controls received drinking water with (SHAM) or without (UNT) LPS. Endotoxin tolerance was tested by determination of LPS-induced tumor necrosis factor (TNF)-alpha release. After the avoidance of the induced endotoxin-tolerance the CS saccharin was re-presented in all experimental groups. A the end of the re-exposure period a complete endotoxin tolerance was noticed in the CONDl- and COND2-group. In contrast, no effect of saccharin administration was observed in the SHAM- or UNT-group. Our data demonstrate for the first time the behavioral conditioning of endotoxin tolerance. Furthermore, these results contribute new aspects to the mechanisms underlying the development and modulation of endotoxin tolerance.     

Intracranial self-stimulation after memory reactivation: immediate and late effects

To assess whether intracranial self-stimulation (SS) given after memory reactivation could improve memory retrieval, we tested the immediate (Experiment 1) and late (24 h; Experiment 2) effects of an SS treatment on the retrieval of a two-way active avoidance conditioning in Wistar rats. Memory was reactivated 24 h after training and the reminder (Rm) used consisted of a 3 s exposure to the conditioned stimulus (a tone) in the same context as in the original learning. SS treatment (2500 trains at 100% of each rat's optimal intensity) was administered immediately afterwards. No significant differences between SS-treated and control groups were observed when the retrieval was tested immediately after the SS treatment with or without memory reactivation. However, retrieval was improved when tested 24 h after SS treatment alone or after the reminder exposure alone. The greatest improvement in avoidance was observed when both treatments were given together, that is, when the SS treatment was administered immediately after memory reactivation. Moreover, there were no significant statistical interactions between the effect of SS treatment and the ones of memory reactivation in any of both experiments. The present results show that the effect of an immediate SS treatment can be added to the ones of memory reactivation causing a strong long-term facilitation of memory retrieval.     

Multiple representations of information in the primary auditory cortex of cats. I. Stability and change in slow components of unit activity after conditioning with a click conditioned stimulus

Recordings of activity were made from 647 single units of the A(I) cortex of awake cats to evaluate behavioral state-dependent changes in the population response to a 70-dB click. Averages of PST histograms of unit activity were used to assess the changes in response. This report focuses on slow components of the responses disclosed by averages employing bin widths of 16 ms. Responses were compared before and after a Pavlovian blink CR was produced by forward pairing of click conditioned stimuli (CSs) with USs. A backward-paired 70-dB hiss was presented as a discriminative stimulus. Studies were also done after backward pairing of the click CSs (backward conditioning) that produced weak sensitization instead of a conditioned response. There were four main findings. First, components of activity elicited 32-160 ms after presenting the hiss decreased significantly after conditioning and after backward conditioning. The decreases after conditioning represented the most pronounced changes in activity evoked by either clicks or hisses in this behavioral state. Second, baseline firing decreased after both conditioning and backward conditioning. The direction of baseline change was opposite that found in adjacent cortical regions and in A(I) cortex after operant conditioning employing an acoustic cue. Third, prior to conditioning, unit activity in response to the hiss declined before the sound of the hiss reached its peak or terminated. This decrease was thought to represent a habituatory adaptation of response to a prolonged acoustic stimulus. This type of habituation to a lengthy stimulus has been recognized, behaviorally, but has not been observed previously in the activity of units of the auditory receptive cortex. Fourth, the percentage of click responsive units did not change significantly after the click was used as a CS for conditioning, and despite the accompanying changes in baseline activity, the absolute levels of activity summed in the first 16 ms after click delivery remained stable across behavioral states in which the motor response to the click was altered profoundly. The onset of the conditioned motor response began 20 ms after the click, and was shown earlier to depend on rapid, potentiated transmission through the cochlear nucleus and motor cortex for its generation. Thus the stability of the response to the click in the primary auditory receptive cortex was unexpected. This led us to make further analyses of the data with 2- and 4-ms bin widths (see companion report) that eventually disclosed a potentiated response to the click. The findings show stability and change in the response to the click as a CS, depending on the band pass (bin width) used for analysis of spike activity. In the representation disclosed by low pass filtering in this study, the response was stable. This representation provided information suitable for identifying commonalties of the click signals across varying behavioral states. The representations of the click and hiss contained in the slow components of the population response in the A(I) cortex were uncorrelated with the selective potentiation of activity in motor cortex and behavioral performance in response to click as a CS after conditioning. Although changes in the activity evoked by hisses occurred after conditioning, the changes also occurred after backward conditioning when only small, sensitized behavioral responses to clicks and hisses were observed. Basic theoretical considerations about information transmission in complex neural networks plus clinical observations comparing derangements of linguistic and non-linguistic cortical functions in humans suggest that multiple representations of conditioned stimulus inputs may exist in local populations of cortical neurons. Together, our studies provide evidence for two different, concurrent representations of information about a click CS encoded in the spike activity of the A(I) cortex.