Gordon A. Barr

Classical conditioning, decay and extinction of cocaine-induced hyperactivity and stereotypy

Following 10 daily pairings of multiple conditioned stimuli with injection of cocaine (15 mg/kg), the presentation of the stimuli alone elicited behaviors in rats similar to those induced by cocaine. The behaviors included increased duration or frequency of rearing, sniffing, head bobbing, and horizontal locomotor activity (crossing). The level of the conditioned response for several of these behaviors approximated that induced by the drug itself. The conditioned drug effect showed decay over 15 days but little extinction during 4 daily trials. Brain concentrations of the dopamine metabolites, homovanillic acid and dihydroxyphenylacetic acid, were similar in the conditioned and pseudoconditioned control groups in both the caudate and mesolimbic areas. The behavioral results demonstrate that, in a classical conditioning paradigm, previously neutral stimuli can elicit behaviors similar to those induced by cocaine and that certain conditioned responses show time related decline. This agrees with the reported conditioning of amphetamines behavioral effects but differs in terms of the action on brain dopamine turnover.

Morphine-induced downregulation of μ-opioid receptors in neonatal rat brain

Chronic administration of morphine to pre- and postnatal rats produced a marked decrease in brain mu-opioid receptor density with-out change in receptor affinity. No significant changes in delta- or kappa-receptors were observed. This downregulation was accompanied by tolerance to the analgesic actions of morphine. In neonates exposed to morphine from postnatal day one, mu-receptor number was significantly depressed until postnatal day 8, then increased gradually to control levels by day 14 of treatment. Longer treatment produced no further change in opioid receptors. These data represent the first demonstration of in vivo downregulation of brain mu-opioid receptors following morphine administration and provide evidence for a unique plasticity of the immature opioid receptor system.

Place conditioning with morphine and phencyclidine: dose dependent effects

In a place conditioning paradigm, rats were exposed to one of two distinctive environments following injection of drug or vehicle. Preference was measured under drug free conditions by allowing subjects free access to both settings and measuring where they spent more time. Comparisons were made between morphine and saline; PCP and saline; and one of several doses of morphine and a standard dose. Morphine was preferred over saline and, when compared to the reference dose, lower doses of morphine were less preferred and higher doses more preferred. PCP was never preferred over saline and under some conditions produced a conditioned place aversion. The ability to generate dose dependent effects with morphine should allow more sophisticated studies in which shifts in dose response curves are required.

Estradiol and progesterone differentially regulate formalin-induced nociception in ovariectomized female rats

Clinical and preclinical studies have found sex-specific differences in the discrimination and perception of inflammatory stimuli. The emerging picture suggests that the biological basis of these differences resides in the regulatory activity of gonadal hormones in the central nervous system. This study describes the effects of ovarian hormones in inflammatory pain processes. Ovariectomized rats received estradiol and/or progesterone, and the number of paw flinches was measured after 1, 2.5 or 5% formalin administration. Both estradiol and progesterone altered the number of flinches only after 1% formalin administration. Estradiol significantly reduced the overall number of flinches during Phase II of the formalin nociceptive response while progesterone attenuated Phase I of the response. After co-administration of estradiol and progesterone, progesterone reversed estradiols analgesic effect in Phase II, however, estradiol did not reverse progesterones analgesic activity in Phase I. To determine if estradiol effects are receptor-mediated, tamoxifen (selective estrogen receptor mediator, 15 mg/kg) or alpha-estradiol (an inactive isomer of estradiol, 20 microg) were utilized. Tamoxifen decreased the number of formalin-induced flinches during Phase II while alpha-estradiol did not affect any formalin-induced responses. When co-administered with estradiol, tamoxifen failed to reverse estradiols effect, suggesting both tamoxifen and estradiol activate similar intracellular mechanisms. Although Western blot analysis detected the presence of estradiol alpha and beta and progesterone B receptors in the spinal cord, hormone replacement treatments had no effects on the levels of these receptors. We postulate that the mechanisms by which estradiol and progesterone induce analgesia occur through the activation of their receptor at the spinal cord level.

κ Opioid receptor-mediated analgesia in the developing rat

The prototypic kappa opiate ketocyclazocine produced robust analgesia in 10-day-old rats in the tail-flick nociceptive test. The kappa-opiate behavioral response coincided with the onset of a rapid rise to adult levels in brain kappa receptor site density. In contrast, morphine (prototypic mu opiate) was without marked effect until 14 days of age. The period of rapid mu receptor increase did not take place until days 14-16, which was after kappa receptor levels had already plateaued. Further, there was no or incomplete cross-tolerance between ketocyclazocine and morphine at 14 days of age. The present study, therefore, establishes a role for the kappa binding site in thermal analgesia in the tail flick test and differentiates its ontogenetic pattern from that of the mu receptor.

Maturation of the biphasic behavioral and heart rate response in the formalin test.

The biological processes that mediate and modulate the perception of pain in the infant animal are not well studied and thus nociception during early development is poorly understood. In the adult animal, injection of formalin into the hind paw produces distinct phases of behavioral and autonomic responses: an early nociceptive response followed by a period of quiescence and a later second phase that matches or exceeds the initial response. The delayed reaction of the second phase has been suggested to be a model of inflammation-induced changes in neuronal sensitivity. Studies in the infant rat have demonstrated that the first phase is present in the fetus and neonate but the onset of the second phase is later maturing. We report here that the first phase occurs in 7- to 35-day-old pups in the formalin test when measured behaviorally and in 14- to 35-day-old pups when assessed by increased heart rate. However, the behavioral response in second phase is greatly attenuated or absent in 7- or 14-day-old pups, a finding consistent with that of others, appearing first at 21 days of age. The biphasic tachycardic response was not noted until even later, at 35 days of age. These data confirm that the neural mechanisms that mediate the secondary behavioral phase in the formalin test are late maturing, that the biphasic cardiovascular response does not occur until substantially later, after weaning, and that the behavioral and cardiovascular responses are dissociated developmentally.

Developmental changes in c-fos expression to an age-specific social stressor in infant rats.

Young rats become immobile when exposed to a potentially infanticidal adult male rat. Male-induced immobility declines during the preweaning period, paralleling the decrease in infanticidal threat. To investigate the neural substrates underlying the developmental change in immobility, male-induced expression of the immediate-early gene c-fos was assessed on postnatal days 7, 14 and 21. A huddle of three young rats was exposed to an adult male behind a screen. As control, three littermates were put in the testing chamber but not exposed to the male. On day 7, male exposed and control pups were immobile most of the time and c-fos expression did not differ between conditions. On day 14, rats in the presence of the male stopped ongoing behaviors and became immobile. They had significantly higher c-fos expression in the paraventricular nucleus of the hypothalamus, the amygdala, the periaqueductal gray, and the locus ceruleus. On day 21, the male-exposed rats that were immobile had elevated c-fos expression in a similar pattern as on day 14, however, different nuclei of the amygdala were activated. In contrast, male-exposed 21-day-old rats that showed control levels of immobility did not have elevated c-fos expression in these areas. These results demonstrate that male exposure induced c-fos expression in brain areas of young rats in an age-specific pattern. Some of the activated brain areas seem to have contributed to immobility. Differential activation of neuronal populations may underlie developmental changes in defensive immobility during early ontogeny.

Morphine- and ketocyclazocine-induced analgesia in the developing rat: differences due to type of noxious stimulus and body topography

Patterns of morphine- and ketocyclazocine-induced analgesia in limb withdrawal and tail-flick tests of thermal and mechanical nociception were examined in the preweanling rat. In the forepaw test, morphine was more effective than ketocyclazocine with both thermal and mechanical stimuli. Both drugs first induced analgesia between 3 and 5 days of age. In the tail-flick test, ketocyclazocine-induced analgesia preceded morphines effects against both thermal and mechanical stimuli by several days. Ketocyclazocine produced robust analgesia between 7 and 10 days of age, while the effects of morphine did not peak until day 14. In the hindpaw, morphine was more effective than ketocyclazocine against a higher intensity mechanical stimulus, while ketocyclazocine was more effective against a lower intensity mechanical stimulus. Morphine-induced analgesia was reversed by lower doses of naloxone than was ketocyclazocine-induced analgesia, regardless of body part tested, against all noxious stimuli. These findings demonstrate differences in morphine- and ketocyclazocine-induced analgesia that are dependent upon age, body topography, stimulus type and intensity and imply different physiologic roles of mu- and chi-opioid receptors in analgesia.

Naltrexone-precipitated morphine withdrawal in infant rat is attenuated by acute administration of NOS inhibitors but not NMDA receptor antagonists

Abstract. Rationale: There is increasing evidence that the N-methyl-d-aspartate (NMDA) receptor and the nitric oxide system are involved in opiate dependence in the adult rat, but whether these results in the adult apply to the infant rat is unknown. Objectives: Here we examined the effects of NMDA receptor antagonists and nitric oxide synthase (NOS) inhibitors, which reduce the opiate abstinence syndrome in adult animals, on morphine withdrawal in the infant rat. Methods: Neonatal rats were injected with morphine sulfate (10.0xa0mg/kg) twice daily for 6.5xa0days. On the 7thxa0day, pups were injected with NOS inhibitors (l-NAME or 7-NI), NMDA receptor antagonists (MK-801 or AP-5), or vehicle. After 15xa0min, the pups were injected with naltrexone (1xa0mg/kg) to precipitate withdrawal. Behavior for each pup was identified and recorded every 15xa0s for 10xa0min before naltrexone injection and 15xa0min after naltrexone injection. Results: Both l-NAME and 7-NI significantly reduced most withdrawal behaviors in the infant rat, a result in line with previous studies in the adult rat. In contrast, AP-5 reduced some withdrawal behaviors but also increased others (e.g., moving paws). MK-801 was likewise ineffective in reducing most withdrawal behaviors and increased certain withdrawal behaviors (walking and wall climbing). Conclusions: In the infant rat, the production of nitric oxide is involved in opiate withdrawal whereas the NMDA receptor may not yet be functionally active or may play only a minor role.

Opiate withdrawal during development: are NMDA receptors indispensable?

Despite decades of research, the mechanisms that underlie opiate tolerance, dependence and withdrawal remain elusive. Evidence accumulated over the past ten years suggests that the NMDA receptor plays a central role in mediating the neuroplasticity induced by chronic opiate administration in adult animals. Yet, during ontogeny, the NMDA receptor complex undergoes qualitative developmental changes, which renders some of the basic assumptions for a role of the NMDA receptor in opiate withdrawal invalid in infants. Recent data indicate that NMDA receptor antagonists are not effective in blocking morphine tolerance, dependence and withdrawal in the neonatal rat. Roles for other glutamate receptor types (e.g. metabotropic glutamate receptors) have also been proposed recently. In this article, the latest evidence that characterizes the dynamic roles of glutamate receptors in these phenomena during ontogeny will be discussed.