Emilio Fdez Espejo

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 Parkinsons 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.

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.

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 rats 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 rats behaviour in the hot plate test following administration of morphine and naloxone has been validated in this study.

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 rats patterns in the hot plate test, as well as to follow the gradual changes in responses when repeated measures are used.

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.

Dorsal and median raphe serotonergic system lesion does not alter the opiate withdrawal syndrome

Previous pharmacological studies have implicated serotonergic brain systems in opiate withdrawal. To test the hypothesis that serotonin (5-HT) has a critical role in the development of opiate withdrawal, we have employed a near-total brain 5-HT system lesion technique (90% depletion) using 5,7-dihydroxytryptamine combined with induction of opiate dependence by implantation of morphine pellets or by repeated injections of increasing doses of morphine. The effects of serotonergic neuron lesion were examined on spontaneous opiate withdrawal (changes in circadian locomotor activity) and naloxone-precipitated opiate withdrawal syndrome (the somatic aspect). The antiwithdrawal properties of clonidine, an alpha(2)-adrenoceptor agonist currently used for clinical treatment for the somatic signs of opiate withdrawal, were tested also in the lesioned rats. Our findings show that serotonergic lesions in morphine-dependent rats did not alter either the spontaneous or the naloxone-induced withdrawal syndrome (with exception of jumping behavior). Moreover, clonidine alleviated the naloxone-induced withdrawal syndrome in lesioned as well as in sham-operated morphine-dependent rats. These results demonstrate that 5-HT systems are not directly responsible for the development of the somatic opiate withdrawal syndrome in morphine-dependent rats.

Changes in dopaminergic neurotransmission do not alter somatic or motivational opiate withdrawal-induced symptoms in rats

Opiate withdrawal has been correlated with decreased extracellular dopamine (DA) levels in the nucleus accumbens (NAC) of morphine-dependent rats. The authors tested the hypothesis that DA transmission plays a critical role in the induction of motivational and somatic withdrawal symptoms. First, the authors used a 6-hydroxydopamine-induced lesion of the NAC to chronically disrupt mesolimbic DA transmission. Second, global DA neurotransmission was acutely stimulated by the nonselective DA agonist (apomorphine) or inhibited by nonselective DA antagonists (droperidol or flupentixol). Morphine-dependent rats bearing 6-hydroxydopamine-induced lesions displayed naloxone-precipitated motivational and somatic withdrawal symptoms similar to those of sham-lesioned rats. Administration of apomorphine did not reduce naloxone-induced opiate withdrawal. Moreover, in total absence of naloxone, DA antagonists did not precipitate either conditioned place aversion or somatic abstinence signs in dependent rats. Taken together, these findings suggested that DA transmission is not critical for the induction of opiate withdrawal syndrome.

Single restraint stress sensitizes acute chewing movements induced by haloperidol, but not if the 5-HT1A agonist 8-OH-DPAT is given prior to stress

The objective of this study was two-fold: (i) to analyze behavioral sensitization to haloperidol 2 weeks after single restraint stress, and (ii) to establish the effects of 8-OH-DPAT treatment prior to stress on sensitized behavioral responses. Overall behavior was analyzed and not only catalepsy, but also sedation (immobility), grooming, exploration and vacuous chewing movements were evaluated. Results indicated that single restraint stress induced a long-lasting sensitization of acute vacuous chewing movements induced by haloperidol (0.25, 0.5 mg/kg i.p.). Interestingly, this behavioral sensitization was prevented by 8-OH-DPAT (0.35 mg/kg s.c.) prior to stress. Finally, haloperidol-induced sedation was not disrupted by either restraint stress or 8-OH-DPAT treatment.

Inhibition of 5-HT neurotransmission increases clonidine protective effects on naloxone-induced conditioned place aversion in morphine-dependent rats.

Previous pharmacological studies have implicated serotonergic brain systems in opiate-withdrawal-precipitated conditioned place aversion. To assess this hypothesis, we tested the effects of either (i) a near-total 5,7-dihydroxytryptamine-induced lesion (90% depletion) or (ii) an acute serotonin (5-HT) inhibition induced by the specific stimulation of 5-HT1A autoreceptors (8-OHDPAT 5–100 μg/kg), on naloxone-induced conditioned place aversion in morphine-dependent rats. Morphine dependence was induced by the implantation of morphine slow-release pellets. The protective properties of clonidine (an alpha-2 adrenergic agonist classically given for opiate detoxification) were also tested after inhibition of 5-HT transmission. Serotonergic lesions in morphine-dependent rats failed to alter naloxone-induced conditioned place aversion but increased the sensitivity to the protective effects of clonidine. Acute neuropharmacological blockade of serotonin transmission also potentiated the clonidine effects on naloxone-induced conditioned place aversion. When combined with the 5-HT1A agonist 8-OHDPAT, clonidine was also found to be more potent. Further understanding of this serotonin/noradrenaline interaction might help devise new therapeutic treatments for the acute opiate withdrawal syndrome.