Functional regeneration in a rat Parkinson's model after intrastriatal grafts of glial cell line-derived neurotrophic factor and transforming growth factor beta1-expressing extra-adrenal chromaffin cells of the Zuckerkandl's organ.

Intrabrain transplantation of chromaffin cell aggregates of the Zuckerkandl's organ, an extra-adrenal paraganglion that has never been tested for antiparkinsonian treatment, induced gradual improvement of functional deficits in parkinsonian rats. These beneficial effects were related to long survival of grafted cells, striatal reinnervation, and enhancement of dopamine levels in grafted striatum. Grafted cells were not dopaminergics, but they expressed glial cell line-derived neurotrophic factor (GDNF) and transforming growth factor-beta(1). These factors were detected in the host striatal tissue, indicating that chromaffin cells secreted them after grafting. Because glial cell line-derived neurotrophic factor possesses neurorestorative properties over dopaminergic neurons, and transforming growth factor-beta(1) is a cofactor that potentiates the neurotrophic actions of GDNF, functional regeneration was likely caused by the chronic trophic action of neurotrophic factors delivered by long-surviving grafted cells. This work should stimulate research on the clinical applicability of transplants of the Zuckerkandl's organ in Parkinson's disease.

Behavioral expression of opiate withdrawal is altered after prefrontocortical dopamine depletion in rats: monoaminergic correlates.

The objective of this study was to establish the effects of prefrontocortical dopamine depletion on opiate withdrawal and prefrontocortical neurochemical changes elicited by morphine dependence and withdrawal. The dopaminergic content was also measured in the nucleus accumbens during withdrawal, in order to detect reactive changes induced by prefrontocortical lesion. Withdrawal was induced by naloxone in morphine-dependent rats. Monoamine levels were analyzed post-mortem by high performance liquid cromatography. The results showed that chronic morphine dependence did not modify basal levels of monoamines in sham rats, revealing neuroadaptation of prefrontocortical dopamine, noradrenaline and serotonin systems to chronic morphine. The neuroadaptive phenomenon remained after prefrontocortical lesion (> 79% dopamine depletion). On the other hand, a strong increase of dopamine, noradrenaline, and serotonin contents in the medial prefrontal cortex of sham rats was detected during opiate withdrawal. However, in lesioned rats, the increase of prefrontocortical dopamine and serotonin content, but not that of noradrenaline, was much lower. In the nucleus accumbens, prefrontocortical lesion reactively enhanced the dopaminergic tone and, although opiate withdrawal reduced dopaminergic activity in both sham and lesioned rats, this reduction was less intense in the latter group. At a behavioral level, some symptoms of physical opiate withdrawal were exacerbated in lesioned rats (writhing, mastication, teeth-chattering, global score) and exploration was reduced. The findings hence indicate that: (i) prefrontocortical monoaminergic changes play a role in the behavioral expression of opiate withdrawal; (ii) the severity of some withdrawal signs are related to the dopaminergic and serotonergic tone of the medial prefrontal cortex rather than to the noradrenergic one, and (iii) an inverse relationship between mesocortical and mesolimbic dopaminergic systems exists.

Adrenergic hyperactivity and metanephrine excess in the nucleus accumbens after prefrontocortical dopamine depletion.

Selective dopamine depletion within the medial prefrontal cortex in rats is known to enhance dopamine and norepinephrine levels in the nucleus accumbens and to induce characteristic behavioral disturbances. The present study was designed to determine levels of adrenaline, apart from dopamine and norepinephrine, and metabolites in the nucleus accumbens after prefrontocortical dopamine depletion. Prefrontocortical dopamine depletion was carried out by injecting 6-hydroxydopamine, and it was validated through: the emergence of behavioral disturbances such as amphetamine-induced stereotypies, spontaneous motor hyperactivity, and enhanced "anxiety-like" responses and through postmortem quantification of catecholamine levels by using high-performance liquid chromatography. The findings indicated that lesioned rats exhibited more oral stereotypies after amphetamine, were hyperlocomotive, and showed more pronounced anxiety-like behaviors than controls. Following prefrontocortical dopamine depletion, postmortem concentrations of dopamine and norepinephrine, along with the metabolites 3,4-dihydroxyphenylacetic acid and vanillylmandelic acid, were reliably enhanced in the nucleus accumbens as expected, and dopamine turnover was decreased. Furthermore the nucleus accumbens contained higher levels of adrenaline and its transmethylated metabolite metanephrine. To sum up, prefrontocortical dopamine depletion induces motor and emotional disturbances in rats and alters the neurochemical profile of the nucleus accumbens, not only inducing dopaminergic and noradrenergic hyperactivity but also leading to adrenaline and metanephrine excess.

Naloxone-induced opiate withdrawal produces long-lasting and context-independent changes in aggressive and social behaviors of postdependent male mice.

The purpose of this study was to determine whether an environment associated with naloxone-induced morphine withdrawal affects aggressive or social behaviors in postdependent mice. Morphine-dependent or saline-treated mice received 3 naloxone injections in 1 of 2 different environments (A or B); 15 days afterward, when the mice were completely drug free, an aggression test was carried out in Environment A. All the mice suffering morphine withdrawal showed a significant increase in aggression, irrespective of the environment in which the withdrawal took place. In these conditions, the impact of morphine dependence and the 3 induced withdrawals was so profound that the environment could not be discriminative. In addition, modifications in the behavioral profile of postdependent mice that suffered only spontaneous withdrawal were long-lasting, with the mice carrying out more attacks during social investigation without presenting threat postures.

Total neurochemical lesion of noradrenergic neurons of the locus ceruleus does not alter either naloxone-precipitated or spontaneous opiate withdrawal nor does it influence ability of clonidine to reverse opiate withdrawal.

It has been suggested that an increase firing rate of noradrenergic neurons of the locus ceruleus is responsible for the opiate withdrawal syndrome. However, lesion studies have indicated that the noradrenergic neurons of the locus ceruleus are not essential for either the expression or suppression by clonidine of opiate withdrawal. The present study was designed to determine the effect of the almost complete 6-hydroxydopamine lesion of noradrenergic neurons (94%) of the locus ceruleus on various components of the opiate withdrawal syndrome and on its protection by clonidine. Morphine dependence was induced by s.c. implantation of morphine pellets (2 x 75 mg base). The following paradigms were used: 1) naloxone-induced conditioned place aversion, 2) naloxone-precipitated acute opiate withdrawal syndrome, 3) nycthemeral locomotor activity as a measure of spontaneous opiate withdrawal. The results showed that quasi-total lesion of noradrenergic neurons of the locus ceruleus did not modify opiate dependence as revealed by naloxone-induced conditioned place aversion and the expression of an acute morphine withdrawal syndrome. Moreover, clonidine prevented the opiate withdrawal syndrome in both lesioned and sham-operated rats, suggesting that the action of clonidine is certainly mediated through postsynaptic alpha(2)-adrenoceptor stimulation. Finally, the nycthemeral locomotor activity during spontaneous morphine withdrawal did not differ between the lesioned and the sham-operated rats.

Prefrontocortical dopamine depletion induces antidepressant-like effects in rats and alters the profile of desipramine during Porsolt's test.

The objective of this study was to investigate whether bilateral dopamine depletion within the medial prefrontal cortex affects depression state, as well as the antidepressant efficacy of desipramine, in the forced swimming test. The rat's behaviour was evaluated by quantifying duration of immobility, climbing, swimming and diving. Immobility latency was also quantified and proved to be a suitable novel parameter. Monoamine levels within the medial prefrontal cortex were measured by high-performance liquid chromatography during Porsolt's test, as well as one week after it. While Porsolt's test was followed by a typical depression-like profile in sham rats, depletion of prefrontocortical dopamine (86% vs sham controls) reduced immobility and enhanced swimming, which is consistent with a diminished depression tonus. The observed enhancement of swimming was correlated with a high prefrontocortical serotonergic neurotransmission. On the other hand, desipramine induced antidepression-like effects in sham rats by increasing prefrontocortical noradrenaline and serotonin neurotransmisson, but also by blocking the normal increase in dopamine activity during the swimming test. Interestingly, desipramine behaved in a quite different manner in lesioned rats. Thus, immobility duration was not further reduced and only climbing, but not swimming, was enhanced. These effects were correlated with a preferential enhancement of noradrenaline neurotransmission. In conclusion, the results indicate that: (i) dopamine neurotransmission within the medial prefrontal cortex is a factor involved in depression, since dopamine reduction led to a low depression tonus; (ii) desipramine induces antidepression not only by enhancing prefrontocortical noradrenaline and serotonin neurotransmission, but also by blocking the normal increase in dopamine neurotransmission during a depressant situation; (iii) a selective enhancement of prefrontocortical serotonin neurotransmission mediates swimming; and (iv) a selectively augmented prefrontocortical noradrenaline activity mediates climbing during Porsolt's test.

Behavioral sensitization to cocaine after a brief social defeat stress: c-fos expression in the PAG.

The experiments explored the nature and time course of changes in behavior and Fos expression in the periaqueductal grey area (PAG) in response to an injection of cocaine that was given following a single episode of social defeat stress. Social defeat stress was defined as an intruder mouse's response to an aggressive resident mouse. First, the intruder was briefly attacked, and secondly, it was threatened while protected by a perforated cage for 20 min. Plasma corticosterone levels rose after the beginning of the confrontation and remained elevated during the protected phase. In a first experiment, separate groups of intruder and control mice were challenged once with cocaine (20, 30, or 40 mg/kg) or saline. During tests for motor activity, behavioral measurements were obtained via (1) photobeam interruptions, (2) tracking of movements via image analysis, and (3) quantitative ethological analysis of postures and acts via videorecords. Several indices of ambulatory or horizontal forward locomotion confirmed the stimulant effects of cocaine. In a further experiment, separate groups of mice were challenged with 40 mg/kg cocaine at one time point, either during the social stress or 3, 5, 7 or 9 days thereafter. A cocaine challenge significantly increased locomotion 5 and 7 days after a brief social defeat stress, in excess of the level that is seen in non-stressed animals. Further experiments used immunohistochemical assays of sections through the caudal ventrolateral PAG and showed a significant increase in Fos-like immunoreactivity (Fos-LI) 1 h after the social stress experience or after cocaine. Importantly, concurrent administration of cocaine with social defeat stress produced inhibition of Fos expression throughout the PAG. A partial to complete recovery of cocaine-induced Fos expression was observed 5-7 days after social defeat stress. The results suggest that a single social stress episode is sufficient to engender a delayed sensitization of stimulant hyperactivity. The initial inhibition of Fos expression by concurrent social stress and cocaine may point to a relevant initiating event in the process of sensitization to stimulants.

Antagonism of peripheral 5-HT4 receptors reduces visceral and cutaneous pain in mice, and induces visceral analgesia after simultaneous inactivation of 5-HT3 receptors.

The role of 5-HT4 receptors on cutaneous and visceral pain remains largely unexplored. The objective of this study was to establish the activity profile of SDZ 205-557, a 5-HT4 antagonist, on cutaneous (hotplate) and visceral (writhing) models of pain, after peripheral administration. Since SDZ 205-557 possesses some affinity for 5-HT3 receptors at high doses, nociceptive effects of a 1:1 combination of SDZ 205-557 and MDL 72222, a 5-HT3 antagonist, were also evaluated. Drugs were injected 30 min before tests (0, 0.001, 0.01, 0.1 or 1 mg/kg IP). A hypoalgesic effect of SDZ 205-557 on cutaneous pain was found at 0.1 and 1 mg/kg doses, as revealed through an enhanced nociceptive threshold in rats placed on the hotplate. This effect was likely mediated through inactivation of peripheral 5-HT4 receptors. After the 1:1 combination, the hypoalgesic effect disappeared, which indicates that simultaneous inactivation of 5-HT3 and 5-HT4 receptors antagonized peripherally 5-HT4-mediated hypoalgesia by an unknown mechanism. SDZ 205-557 also induced hypoalgesia in the writhing test over the entire dose range tested, and visceral hypoalgesia turned out to be analgesia after 1:1 combination. In summary, findings of the present study imply that: i) antagonism of 5-HT4 receptors mediates antinociception in enteric viscera and, to a lesser extent, in cutaneous terminals, and ii) dual inactivation of both 5-HT4 and 5-HT3 receptors induces visceral analgesia, a fact which might have clinical importance.

Cellular and functional recovery of Parkinsonian rats after intrastriatal transplantation of carotid body cell aggregates.

We have tested the suitability of chromaffin-like carotid body glomus cells for dopamine cell replacement in Parkinsonian rats. Intrastriatal grafting of cell aggregates resulted in almost optimal abolishment of motor asymmetries and deficits of sensorimotor orientation. Recovery of transplanted animals was apparent 10 days after surgery and progressed throughout the 3 months of the study. The behavioral effects were correlated with the long survival of glomus cells in the host brain. In host tissue, glomus cells were organized into glomerulus-like structures and retained the ability to secrete dopamine. Several weeks after transplantation, dopaminergic fibers emerged from the graft, reinnervating the striatal gray matter. The special durability of grafted glomus cells in the conditions of brain parenchyma could be related to their sensitivity to hypoxia, which is known to induce cell growth, excitability, and dopamine synthesis. This work should stimulate research on the clinical applicability of carotid body autotransplants in Parkinson's disease.

Effects of weekly or daily exposure to the elevated plus-maze in male mice.

The elevated plus-maze is an animal model where the behavioural repertoire of rodents is used to detect effects on anxiety. Repeated testing is a procedural variable where contradictory results have been reported. Some laboratories have found stable test-retest profiles, although other studies have reported reduced open arm exploration. The objective was to further discern behavioural changes in the behaviour of the mouse after either weekly or daily tests. Behaviour was videotaped and later analysed. Behavioural patterns were encoded from an ethological point of view, a nine-pattern ethogram being employed. Other parameters such as percent time on the different sections of the maze were evaluated as well. Descriptive analysis revealed a progressive decrease in percent time spent on open arms (in weekly-tested mice), percent time on central platform, open arm entries, percent open entries, unprotected stretched attention posture (uSAP) and unprotected head-dipping (uDip), together with an augmented number of closed arm returns and percent time spent on closed arms. Taken together, these findings are consistent with an enhanced anxiety level across the tests. It is worth noting that percent time on open arms, a traditional anxiety-related parameter, was not progressively decreased in daily-tested mice. Other than expected, exploratory and locomotor elements such as sniffing, rearing, closed arm entries, and total arm entries remained quite similarly elicited throughout the tests, suggesting that locomotor habituation was not developed. However, grooming, considered a displacement response, habituated across the tests. In conclusion, the findings of the present study support the hypothesis that anxiety is enhanced after test repetition, and indicate that test-retest profiles are far from stable, except for exploratory locomotor activity.

Selective dopamine depletion within the medial prefrontal cortex induces anxiogenic-like effects in rats placed on the elevated plus maze.

The objective of this study was to investigate if selective dopamine depletion within the medial prefrontal cortex modifies the anxiety state in rats. Anxiety was evaluated by using the elevated plus maze test, an anxiety model. Dopamine depletion in the medial prefrontal cortex (79% vs. controls) induced a significantly lower preference to stay on open arms together with a reliably lower frequency of open arm entries, as well as a significant increase of percent time spent on closed arms. Although locomotion was also significantly reduced, protected head-dipping and protected stretched attend, novel "ethologically derived" indices of anxiety, were reliably enhanced. Taken together, the results are indicative of enhanced anxiety level despite hypomotility. The findings confirm that prefrontocortical dopamine activation is necessary for coping with an anxiogenic challenge, allowing the animal to display adaptive exploratory responses in a fear-inducing environment.

Differential effects of weekly and daily exposure to the hot plate on the rat's behavior.

Animals were assigned to two groups: weekly stimulated (WS, n = 30) and daily stimulated (DS, n = 30). Three hot plate tests (55.0 +/- 0.5 degree C, 45 s exposure time) were carried out for each rat. Behavior was videotaped and analyzed by a 14-pattern ethogram and a software package. A cluster analysis revealed that naive rats mainly displayed: i) exploratory patterns (walk-sniff, immobile-sniff), ii) primary noxious-evoked elements (forepaw licking, hindpaw licking, stamping), iii) escape responses (learning posture, jumping), and iv) hindleg withdrawal, an independent noxious-evoked element. The main behavioral changes over time were: i) exploratory behavior was habituated in both groups, ii) sensitization of primary noxious-evoked elements was observed only in the WS group, iii) escape elements were enhanced in both groups, although to less extent in the DS group, and iv) hindleg withdrawal was enhanced in both groups. Furthermore, in the WS group, the nociceptive threshold was significantly decreased, and cluster analysis indicated reliable changes over time. Results suggest that a sensitization phenomenon came about when rats were tested weekly, but was minimized by using the daily testing schedule. This study indicated that an ethological analysis is useful to categorize the rat's patterns in the hot plate test, as well as to follow the gradual changes in responses when repeated measures are used.

Effects of morphine and naloxone on behaviour in the hot plate test: an ethopharmacological study in the rat.

The objectives of this study were: i) to analyse the effects of morphine and naloxone on the rat's behaviour in the hot plate test using an ethological approach, and ii) to compare the effectiveness of repeated versus single test paradigms. Animals received either morphine (0, 3, 6 or 9 mg/kg SC) or naloxone (0, 0.01, 0.1 or 1 mg/kg SC). For repeated hot plate measures, rats were tested before and 60, 120, 180 and 240 min following morphine treatment, as well as 30, 60, 90 and 120 min after naloxone injection. For the single test schedule, rats were tested only once 60 min after morphine or 30 min after naloxone administration, or at 60, 120, 180, 240 and 300 min after 9 mg/kg morphine treatment. Behaviour was videotaped and analysed by an ethogram and ethological techniques. A cluster analysis revealed that the most frequently displayed patterns could be categorised into exploratory sniffing reactions (walk-sniff, immobile-sniff) and noxious-evoked elements, including primary (paw-licking, stamping), escape (jumping, leaning posture) and independent (hindleg-withdrawal) patterns. During repeated tests, morphine treatment induced: i) a maximum hypoalgesic effect 60 min post-injection (noxious-evoked patterns were significantly reduced), and ii) an unexpected "thermal hyperreactivity rebound effect" after 120 min (paw-licking and hindleg-withdrawal were enhanced), although changes in hindpaw-licking are more indicative of a hyperalgesic rebound effect. Most changes were quite similar during the single test schedule at 60 and 120 min after morphine injection. With regard to naloxone treatment, jumping latency was significantly decreased during the repeated test schedule, but not on single exposure to the plate. Other elements were facilitated, however, in the single test (stamping, leaning posture, hindleg-withdrawal). The results indicated that both repeated and single tests paradigms are of value for testing the effects of morphine and naloxone on rats. However, under our conditions the single test paradigm gave a better picture of the overall effects of the drug. Learning as well as habituation and sensitization may mask certain effects during repeated tests. In conclusion, an ethological analysis of the rat's behaviour in the hot plate test following administration of morphine and naloxone has been validated in this study.

Structure of the rat's behaviour in the hot plate test.

The objectives of the study were: (i) to elaborate an ethogram of the rat's behaviour during the hot plate test, (ii) to analyse the rat's responses to the nociceptive heat stimulus by using ethological techniques, and (iii) to discern the suitable pattern or patterns to evaluate the rat's reaction to the noxious stimulus. Male Wistar rats (n = 30) were employed, and one hot plate test was carried out for each rat. The platform (Socrel DS37) was maintained at 55.0 +/- 0.5 degrees C, and the exposure time was 45 s. Behaviour was videotaped and analysed by an ethogram and a software package. Every pattern parameter (frequency, duration and latency for the first occurrence) was quantified. Cluster and Markovian analyses were also employed, whereby a dendrogram and a Markov chain were obtained, respectively. The ethogram was composed of twelve patterns: immobile-sniff, walk-sniff, rearing, forepaw-licking, hindpaw-licking, face-washing, body-cleaning, stamping, hindleg-withdrawal, leaning posture, jumping off, and freezing. Cluster analysis indicated that the most frequent patterns could be classified into three categories: sniffing responses (immobile-sniff, walk-sniff), primary noxious-evoked patterns (forepaw-licking, hindpaw-licking, stamping), and escape noxious-evoked responses (leaning posture, jumping off). Hindleg-withdrawal would be an independent noxious-evoked pattern, and freezing an alerting behaviour. The ethological information could be useful to better elucidate effects on rats' behaviour of drug treatment or other phenomena during the hot plate test.