Elaboration of a conditioned reflex in a single experiment with simultaneous recording of neural activity.

In 18 of 20 experiments with click CS, and in 5 of 10 experiments with flash CS, we have elaborated a conditioned EMG response in a single session (less than 60 pairings). The US was direct electrical stimulation of the rabbit's motor cortex that evoked a movement of the forelimb. Electrical stimulation of the lateral hypothalamus was used as reinforcement. Parameters for this reinforcement were chosen to evoke a feeding reaction or self-stimulation in the freely behaving animal. The elaborated EMG response satisfied most of the required characteristics of a conditioned reflex. These included spontaneous recovery after extinction, savings of long duration, specificity to stimulus pairing, and weak efferent and afferent generalization. In most experiments with click CS, the elaborated phasic response had an amplitude of 0.1-6 mV with a latency of 12-16 msec. In form and latency the conditioned response was similar to the unconditioned startle reaction of the same animal under chloralose anesthesia, or to its unanesthetized response to a loud sound. However, the conditioned response differed from the startle reaction in that it was localized. Extracellular recordings of 2-5 neurons were simultaneously made from sensory motor cortex near the point at which the US was applied. In 7 experiments 17 neurons were followed through the entire cycle of elaboration and extinction of the conditioned response. Seven neurons showed a statistically significant (P less than 0.05) increase of the response to CS during conditioning. Latencies were 20-140 msec. Interactions between neurons were studied by computing cross-correlograms and joint PST scatter diagrams. These measures were less informative than we had hoped because of the low level of spontaneous and evoked activity, and because of the small numbers of stimulus presentations that were needed for elaboration and extinction of the conditioned reflex. In isolated presentations of the CS after elaboration of the CR, we sometimes observed neural responses to click with a latency less than 6 msec. We propose that at least one of the pathways involved in the localized conditioned startle reflex reported here goes through the sensory motor cortex.     

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.     

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.     

Amygdaloid central nucleus neuronal activity accompanying pavlovian cardiac conditioning: effects of naloxone

Rabbits were treated intravenously with either naloxone-HCl (0.5 mg/kg) or saline vehicle prior to aversive pavlovian conditioning and extinction training; heart rate conditioned responses and concomitant multiple-unit activity in the amygdaloid central nucleus were compared. Multiple-unit activity evoked by the conditioned stimulus increased during conditioning and decreased during extinction in saline-treated rabbits; naloxone treatment attenuated evoked neuronal activity but enhanced bradycardiac conditioned responses. Correlational analysis showed that, in 3 of 8 animals in the saline-treated group, larger increases in multiple-unit activity corresponded to smaller bradycardiac responses. Naloxone treatment did not alter the frequency or magnitude of this relationship, but it did augment the training-induced decrements in evoked neuronal activity at placements that were correlated with bradycardiac response magnitude. These data suggest that conditioned decreases in central nucleus neuronal activity normally may serve to disinhibit vagal mechanisms as conditioned bradycardia develops and that the neural circuits that produce these responses are sensitive to opioid modulation.     

Cardiovascular and behavioural components of conditioned fear to context after ganglionic and alpha-adrenergic blockade

This study investigates the contribution of the peripheral nervous system to the cardiovascular component of long lasting (40 min) conditioned fear responses to context. The conditioned fear response evoked by reexposure to a footshock chamber was tested 10 min after intravenous injection of either the nicotinic ganglion blocker chlorisondamine (0.6 mg/kg) or the alpha-adrenoceptor antagonist phentolamine (10 mg/kg) in six rats implanted with telemetric probes. Compared to saline controls, chlorisondamine did not change the behavioural component of the response (freezing, ultrasonic vocalizations) but almost completely abolished its cardiovascular component (mean arterial pressure and heart rate). Phentolamine also abolished the pressor response but increased the cardiac response, and ultrasonic vocalizations were reduced by half. The results indicate that the long lasting pressor response of conditioned fear to context is sympathetically mediated like the much shorter pressor response of conditioned fear to a discrete stimulus.     

EFFECTS OF PSYCHOTROPIC DRUGS ON BEHAVIOR AND EEG, FOLLOWING DIRECT ADMINISTATION TO DOG'S BRAIN

1. Two kinds of psychotropic drugs, chlorpromazine and imipramine, were examined in forty-one dogs by means of the conditioned avoidance response and the electrocorticogram. These drugs were administered through three routes, namely intra-peritoneal, intra-arachnoid, and intra-ventricular injection, with neither anaesthesia nor restraint. 2. Following the intra-arachnoid injection: Chlorpromazine produced the slow conditioned avoidance response in behavior, and asymmetrical ECoG, synchronized pattern. Imipramine demonstrated no attenuation of the conditioned avoidance response, and slight synchronization on the ECoG. The depressant action of chlorpromazine to the cerebral cortex was confirmed. 3. Following the intra-ventricular injection: Both drugs blocked tentatively the conditioned avoidance response and produced a continuous desynchronized pattern on the ECoG. The dissociation between the behavior and the ECoG was noticeable. The periventricular area might have a more important role on the conditioned avoidance response than the cerebral cortex. 4. The difference of drug action between chlorpromazine and imipramine was manifested following the intra-arachnoid injection. 5. The significance of the function of the cerebral cortex and periventricular structures to the conditioned avoidance response and to the action of the drug was discussed.     

Lesions of the amygdaloid central nucleus block conditioned cardiac arrhythmias in the rabbit receiving digitalis

We have previously reported [15] that malignant cardiac arrhythmias can be evoked in the rabbit receiving digitalis by cardiovascular changes in response to (a) a Pavlovian aversively conditioned stimulus (CS) or (b) electrical stimulation of the amygdaloid central nucleus (ACE), a structure which contributes to Pavlovian-conditioned cardiovascular responses in the rabbit. The present study was designed to examine further the role of the ACE in arrhythmogenesis by determining the effect of lesions of the ACE on the occurrence of CS-evoked arrhythmias during digitalis administration in the rabbit. Two groups of rabbits (ACE lesion and SHAM) received electrodes implanted bilaterally in the ACE, while a third group (UNOP) served as an unoperated control. All rabbits were given Pavlovian conditioning trials in which a tone conditioned stimulus (CS) was paired with an eyelid shock as an unconditioned stimulus (US). Twenty-four h later, rabbits in the lesion group received bilateral lesions of the ACE. Twenty-four h after the lesion, rabbits in all three groups were given a retention test in which an i.v. infusion of ouabain was delivered, followed by 20 CS alone trials. Presence or absence of arrhythmias was scored during the pre-CS baseline and CS periods for each trial. All three groups exhibited few instances of pre-CS baseline arrhythmias, the frequency of which did not differ between groups. The SHAM and UNOP control groups exhibited a significant increase in the occurrence of CS-arrhythmias compared to pre-CS levels. This increase was blocked in the group receiving lesions of the ACE, as was CS-induced bradycardia which typically occurs in response to the CS in the rabbit.(ABSTRACT TRUNCATED AT 250 WORDS)     

The conditioned enhancement of neutrophil activity is catecholamine dependent

Neutrophil activity was elevated in the conditioned mice for the first time through an established conditioned training process. Catecholamines were proved to be important in the regulation of this conditioned innate immunity. In the study, the camphor odor (as the conditioned stimulus, CS) and poly I: C (as the unconditioned stimulus, US) was used to conditionally elevate the activity of the splenic neutrophils. The mechanism(s) responsible for the conditioned enhancement of neutrophil activity was further investigated using the neurochemical blocking assay and immunohistochemical analysis. Results showed that the neutrophil activity was significantly enhanced through the conditioned training process; both reserpine and 6-hydroxydopamine (6-OHDA) significantly blocked this conditioned innate immunity at the conditioned recall stage. Dexamethasone (Dex), however, showed no effect on the conditioned neutrophil response. Tyrosine hydroxylase (TH)-positive cells significantly increased in the locus coeruleus (LC), hypothalamus, and cortex but not in the spleen of the conditioned animals. These results indicate that during the conditioned recall stage, the brain signals the splenic neutrophils via the sympathetic nervous system (SNS) by releasing the peripheral catecholamines in spleen. The activation of the SNS, on the other hand, is also under the influence of catecholamines released in the LC. The hypothalamic pituitary (HP) axis, on the other hand, plays no role in the regulation of the conditioned neutrophil response.     

Evoked potential conditioning using morphine as the unconditioned stimulus in rats

It is believed that conditioning processes play an important role in the development of narcotic dependence and may be particularly responsible for relapse after treatment. In addition, it was suggested that this conditioning is reflected by altered neural responses to stimuli which were associated with drug administration. To test these hypotheses, evoked potentials were recorded from four regions of rats' brains during a differential classical conditioning procedure. The conditioned and unconditioned effects of morphine on neural responses in each of these regions were studied during both addiction and withdrawal. The conditioning procedure resulted in alterations in the cortical response to the stimulus which was associated with morphine, but no alterations occurred in the response to the differential stimulus. Furthermore, the conditioned cortical response was found to persist after withdrawal from morphine. However, possibly due to state-dependency effects, the conditioned response was expressed only when the subjects were morphine intoxicated. Thus, the conditioned neural response found in this experiment did not reflect the processes postulated to lead to relapse of drug-free addicts.     

Operant conditioning of cortical visual evoked potentials in the curarized rat and control of simultaneous subcortical activity

Curarized rats were successfully trained to modify the amplitude of a certain portion of their cortical visual evoked potential. The positive results obtained enabled us to conclude that the influence of peripheral mediators on the generation of the conditioned response is highly improbable. Subcortical visual evoked potentials were simultaneously recorded in the lateral geniculate body, mesencephalic reticular formation, and superior colliculus. As the responses were almost identical throughout the entire experimental session, those structures cannot be held responsible for the conditioned modifications of the cortical evoked response. On the contrary, the nucleus posterior thalami responses paralleled the cortical modifications. But electrolytic lesions of this nucleus proved to have no effect on the evolution of the conditioning procedure. In the rat, the existence of a projection arising from the visual cortex to the nucleus posterior thalami is considered.     

Conditioned locomotion induced by unilateral intrastriatal administration of apomorphine: D(2) receptor activation is critical but not the expression of the unconditioned response

The present study examined the role of D(1) and D(2) receptors in the conditioning of apomorphine-induced locomotor behavior. A Pavlovian conditioning protocol was used in which rats received 5 daily intrastriatal apomorphine treatments paired or unpaired to an open-field environment followed, 2 days later, by a saline test for conditioning. In the conditioning induction phase, the intrastriatal apomorphine treatment increased locomotor activity expressed as an increased number of sectional crossings and rearings. In the conditioning test, the apomorphine-paired group had significantly higher locomotor activity than the unpaired and vehicle groups, consistent with the development of a conditioned locomotor response. The concomitant blockade of D(1) and D(2) receptors with D(1) (SCH23390) and D(2) (sulpiride) antagonists prevented the apomorphine-induced behavioral response during the induction phase and in the conditioning test. Pretreatment with the D(1) antagonist SCH 23390 also blocked the apomorphine-induced behavioral response during the induction phase but did not block the apomorphine conditioned response. Pretreatment with the selective D(2) antagonist sulpiride blocked the apomorphine behavioral response during the induction phase and in the conditioning test. Altogether, these results indicate that antagonism of either the D(1) or D(2) receptors in the dorsal striatum can block apomorphine-induced locomotor activation but that D(2) but not D(1) antagonism can prevent the development of the apomorphine conditioned response. Altogether, these findings indicate a key role for the D(2) receptor site in the mediation of apomorphine conditioned behavior; and, in addition, that apomorphine conditioned locomotor response can develop without the expression of the locomotor stimulant response during the induction phase of conditioning.     

The involvement of glutamate in recall of the conditioned NK cell response

The molecular mechanisms responsible for the conditioned enhancement of natural killer (NK) cell activity were investigated. The primary goal of the study was to examine the roles of glutamate and gamma-aminobutyric acid (GABA) in recall of the conditioned NK cell response. Both neurochemical blocking assay and high performance liquid chromatography (HPLC) technique were used in the study. Results from the neurochemical blocking assay demonstrated that glutamate but not GABA was required in recall of the conditioned NK cell response. NMDA but not the kainate/AMPA receptors, are believed to be involved. The levels of glutamate that were released and/or taken up also appeared to be critical in that interruption of glutamate release and/or uptake blocked the conditioned NK cell response. Results from the HPLC analysis, however, did not show any significant difference in the glutamate content between the conditioned and control brains.     

Conditioned fear to environmental context: cardiovascular and behavioral components in the rat

This study compares the time course of the cardiovascular changes (mean arterial blood pressure, heart rate) and behavioral changes (freezing, rearing, grooming and activity) evoked by 30 min long exposures to a footshock chamber before and after conditioning with footshocks. The main finding is that the conditioned fear evoked by re-exposure to the footshock chamber after conditioning is associated with a prolonged freezing response, a marked rise in mean arterial pressure (+35 mm Hg above a resting baseline of 105 mm Hg) and a delayed rise in heart rate. The pattern of behavioral and cardiovascular changes is the same as with conditioned fear to a discrete stimulus but the effect is a lot longer.     

Circumvention of extraretinal photoresponses in assessing recovery of vision following optic nerve crush in goldfish

Earlier experiments indicated that a conditioned light stimulus that was used to investigate the recovery of vision following optic nerve crush could evoke an extraretinal photoresponse. The present experiments sought to identify a visual stimulus that does not evoke a response following removal of both eyes for use in experiments on optic nerve regeneration. A stimulus consisting of a slight up-down movement of a small ring of light, that was kept stationary between conditioning trials, was classically conditioned to shock in eyed fish and the conditioned response consisted of a suppression of branchial ventilation movements. Following bilateral enucleation or a sham operation the fish received additional sessions of conditioning trials over a period of 3 weeks. Postoperative responding to the moving light stimulus was blocked in the enucleated but not the sham-operated fish. When the ring of light was turned on and off as a conditioned stimulus, responding was extensively but not completely eliminated following enucleation. The investigation confirms that extraretinal photostimulation can be classically conditioned to shock in at least some goldfish, and it shows that such conditioning can be circumvented by using a small moving light as the conditioned stimulus.     

Naltrexone blocks the expression of the conditioned elevation of natural killer cell activity in BALB/c mice

An elevation of natural killer (NK) cell activity was conditioned by the association of a camphor odor conditioning stimulus (CS) with an injection of 20 micrograms polyinosinic:poly-cytidylic acid (poly I:C), the unconditioned stimulus (US). Poly I:C elicits the production and secretion of interferon (IFN), which induces an increase in NK cell activity. Reexposure to the CS occurred on Days 3 and 5 after the association trial on Day 0. Immediately following the CS exposure on Day 5, 1 microgram poly I:C was administered to all animals. This procedure resulted in an increased NK cell activity in the conditioned (CND), but not the nonconditioned (NC), mice. In this study we have shown that the expression of the conditioned response was blocked by an injection of naltrexone (NTX) at 10 mg/kg ip when given immediately prior to the two test CS odor exposures. Peripheral treatment (ip) with a quaternary form of naltrexone (QNTX), which is a less potent opiate antagonist, at the same dose and at the same time relative to the CS odor reexposure did not block the conditioned response. The formation of the conditioned association did not appear to be disrupted by NTX at the 10 mg/kg dose when given immediately prior to the trial odor exposure on Day 0. No modulation of NK cell activity was observed in any of the control groups treated with naltrexone or the quaternary analog. Because of the inability of the QNTX to block the conditioned response, we hypothesize that the opiate receptors involved in the conditioned response and blocked by NTX were within the central nervous system (CNS). Whether this response is peripherally or centrally mediated, we have shown that opiate receptors represent part of the mechanism which mediates the conditioned augmentation of NK cell activity.     

Effects of Noradrenaline on Frequency Tuning of Rat Auditory Cortex Neurons

The selectivity of rat auditory cortex neurons for pure tone frequency was studied during and after ionophoretic application (5–40 nA) of noradrenaline in urethane-anaesthetized rats. The dominant effect induced by noradrenaline was a significant decrease in spontaneous (93/268 cells) and evoked activity (1331268 cells) which outlasted the application. In the whole population of cells (n = 268) the signal-to-noise ratio, computed using as the signal either the mean evoked response or the response at the best frequency, was unchanged during noradrenaline application. It was significantly increased only for cells showing significantly decreased spontaneous activity, and was significantly decreased for cells showing increased spontaneous activity. Frequency selectivity was significantly increased for the whole population during and after noradrenaline application. It was also significantly increased for cells showing significantly decreased evoked activity, and was significantly decreased for cells showing increased evoked activity. The noradrenaline-induced inhibition was not blocked by propranolol (β antagonist); it was blocked by prazosin (α1 antagonist) and partly mimicked by phenylephrine (α1 agonist). GABA, which also inhibited spontaneous and evoked activity, slightly increased the signal-to-noise ratio and significant increased frequency selectivity. However, when noradrenaline was ejected in the presence of bicuculline at doses that were able to block GABAergic inhibition, the inhibitory effects of noradrenaline on spontaneous and evoked activity were still observed. The possible function of noradrenaline-induced inhibitions in sensory cortices is briefly discussed.     

A supratrigeminal region implicated in the classically conditioned nictitating membrane response

The dorsolateral pontine brain stem was investigated as a possible locus of neural elements mediating the classically conditioned nictitating membrane (NM) response in rabbit. Recording and brain stimulation were employed for this purpose. Low-impedance tungsten monopolar microelectrodes were chronically implanted into the pontine brain stem. Multiple-unit recording during classical conditioning revealed a conditioned increase in multiple-unit activity (MUA) which developed and extinguished concurrently with the acquisition and extinction of the behavioral conditioned response. Pseudoconditioning and conditioned inhibition controls indicated that the increase in MUA was an associative learning phenomenon. Histology indicated that electrode tips recording the CR-associated electrical activity were located mostly adjacent or dorsal to the motor trigeminal nucleus. Periocular shock pulses elicited short latency evoked responses throughout most of the dorsolateral pons, suggesting that information concerning the unconditioned stimulus is relayed to this region. Furthermore, electrical stimulation of this region produced a robust ipsilateral nictitating membrane response in a number of cases, suggesting that neural elements of the dorsolateral pons project to the motoneurons that produce membrane extension. A consideration of several criteria based on these experiments implicates a supratrigeminal zone [22] as containing the neural elements of dorsolateral pons most intimately associated with the conditioned NM response. Other interpretations, concerning fibers of passage through this region and its possible relationship to cerebellum [17] are discussed.     

Conditioned lick behavior and evoked responses using whisker twitches in head restrained rats

To examine whisker barrel evoked response potentials in chronically implanted rats during behavioral learning with very fast response times, rats must be calm while immobilized with their head restrained. We quantified their behaviors during training with an ethogram and measured each individual animals' progress over the training period. Once calm under restraint, rats were conditioned to differentiate between a reward and control whisker twitch, then provide a lick response when presented with the correct stimulus, rewarded by a drop of water. Rats produced the correct licking response (after reward whisker twitch), and learned not to lick after a control whisker was twitched. By implementing a high-density 64-channel electrocorticogram (ECoG) electrode array, we mapped the barrel field of the somatosensory cortex with high spatial and temporal resolution during conditioned lick behaviors. In agreement with previous reports, we observe a larger evoked response after training, probably related to mechanisms of cortical plasticity.     

Effect of 5,7-dihydroxytryptamine on the food-aversive conditioning in the snail Helix lucorum L

The effects of 5,7-dihydroxytryptamine (5,7-DHT), a drug which selectively ablates serotonergic terminals, were examined on acquisition of food-aversive conditioned reflex in the snail Helix lucorum. The percent of feeding reactions decreased from 80 to 15% in the conditioned group of animals after 5-8 pairings of food and electric shock. The behavioral performance of 5,7-DHT-injected animals after the same training session coincided with the data received from the unpaired control group: the percent of feeding reactions remained the same as before the training. Conditioning was carried out on the semi-intact 'lip-CNS' preparations as well. Intracellular recordings from the neurons responding to the withdrawal reaction confirmed the results of the behavioral experiments. Elaboration of associative changes was effective on preparations made from normal snails, whereas no changes were noted in 5,7-DHT-treated and pseudoconditioned animals. In 5,7-DHT-treated animals some components of the feeding behavior and withdrawal reaction changed as well. The appetitive phase duration of feeding lengthened significantly, moreover the sensitization of the withdrawal reaction evoked by rhythmic tactile stimulation disappeared in preparations made from drug-treated snails.     

Dopamine receptors mediate drug-induced but not Pavlovian conditioned contralateral rotation in the unilateral 6-OHDA animal model

Following Pavlovian conditioning treatment sessions with apomorphine, animals receiving the paired treatment showed substantial contralateral rotation when placed without drug into the test environment previously paired to the apomorphine (0.5 mg/kg) injection while animals in the unpaired control treatment showed only ipsilateral rotation. Subsequent tests with the D1 antagonist (SCH 23390), or the D2 antagonist (haloperidol) partially suppressed and the combined D1-D2 antagonists completely suppressed the apomorphine-induced response of contralateral rotation. The identical contralateral rotation response occurring as a Pavlovian conditioned response in the paired apomorphine treatment group was not attenuated by dopamine receptor blockade. In both paired and unpaired groups, the spontaneous ipsilateral rotation response was completely blocked. Thus, non-dopaminergic mechanisms mediate conditioned rotation whereas the drug-induced as well as the spontaneous rotation responses require stimulation of striatal dopamine receptors.