Richard M. Kream

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

Abstract 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.

Altered tachykinin expression by dorsal root ganglion neurons in a rat model of neuropathic pain

&NA; The experiments described in the present study approached nerve injury from both a biochemical and anatomical perspective by monitoring changes in expression of preprotachykinin (PPT) mRNA encoding the prototypic tachykinin substance P and related peptide species in neurons of the rat dorsal root ganglia (DRG) following unilateral chronic constriction injury of the sciatic nerve. In situ hybridization histochemistry (ISHH) analyses in conjunction with computer‐assisted image processing were employed to quantify levels of PPT mRNA distributed in DRG neurons. Injury‐induced changes in PPT mRNA expression by affected DRG neurons included:(1) at early postoperative times, generally increased levels of PPT mRNA associated with small and intermediate‐size B cells exhibiting normal morphology,(2) at late postoperative times, markedly decreased levels of PPT mRNA associated with degenerating B cells, and(3) induction of PPT gene expression by large A cells which is highly correlated with degenerative morphological changes. The significant aspects of these changes are discussed with special emphasis on the contribution of altered transmitter expression by DRG neurons to the pathophysiology of causalgia. In particular, the induction of PPT gene expression by many of the large neurons undergoing degenerative changes may represent an important biochemical parameter which is associated with the development and persistence of experimental allodynia.

Effect of early exposure to δ-9-tetrahydrocannabinol on the levels of opioid peptides, gonadotropin-releasing hormone and substance P in the adult male rat brain

The effects of neonatal exposure to delta-9-tetrahydrocannabinol (THC) on the adult animal brain neurochemistry and pain perception were evaluated. Newborn rat pups were culled to a litter size of 8 (males and females) and treated either with THC (2 mg/kg) or oil (control) daily, during days 1-4 after birth. After weaning, the THC-treated males were housed 4 per cage. During the juvenile period (day 50), the THC-treated animals exhibited significantly lower baseline tail-flick values (a measure of pain perception) than the control. However, as adults, the THC-treated animals exhibited significantly higher sensitivity to pain following 5 mg/kg morphine challenge. Furthermore, the THC-treated animals had significantly elevated beta-endorphin and methionine-enkephalin levels in almost all the brain areas sampled for the study. In addition, the neonatally THC-treated rats exhibited significantly higher levels of substance P (SP) and significantly lower levels of gonadotropin releasing hormone (GnRH) in the anterior hypothalamus-preoptic area. The SP and GnRH levels did not differ among the THC-treated and control animals in the medial basal hypothalamus. The results of this study indicate that even a very low dose of THC administered during the neonatal period has a long-lasting effect on the brain neurochemistry. In particular, neonatal administration of THC appears to alter functioning of the endogenous opioid system.

The effect of diet on cribbing behavior and plasma β-endorphin in horses

Five cribbing horses and six control horses were used in a latin square design dietary study to investigate the effects of different diets on the frequency of cribbing behavior and plasma levels of beta-endorphin in the horse. Feeding grain or sweetened grain rations was found to cause a significant increase in the cribbing frequency whereas alfalfa pelleted hay was without significant effect on the frequency of the behavior. Baseline beta-endorphin levels in cribbing horses were half those of the non-cribbing controls and remained signif- icantly lower during the feeding trials. These results are discussed as they apply to treat- ment of cribbing horses and in terms of the underlying mechanism of cribbing.

Sufentanil, morphine, met-enkephalin, and kappa-agonist (U-50,488H) inhibit substance P release from primary sensory neurons: a model for presynaptic spinal opioid actions.

An in vitro model system for analysis of presynaptic inhibitory actions of spinal opioids has been applied. Embryonic sensory neurons derived from chick dorsal root ganglia were grown in primary cell culture, and the release of substance P was evoked by electrical field stimulation during exposure to drugs with well-demonstrated affinity for opioid receptors. This allowed a pharmacologic characterization of the inhibitory actions of specific opioid agonists on the release of substance P as measured by radioimmunoassay (RIA). Sufentanil (0.5 microM), a high affinity mu receptor agonist, U-50,488H (25 microM), a selective kappa receptor agonist, and morphine (10 microM), an agonist with high affinity for mu and delta receptors, inhibited the evoked release of substance P by approximately 60%, 40%, and 50%, respectively. For sufentanil the response was demonstrated to be dose-dependent. As is the case for its analgesic action in vivo, morphine was approximately 50-fold less potent than sufentanil on a molar basis in this assay. The actions of sufentanil, U-50-488H and morphine were mimicked by the endogenous opioid peptide met-enkephalin, and its stable synthetic analog D-ala2-met5-enkephalinamide (DAME). Naloxone (25 microM), an opioid receptor antagonist, blocked the inhibitory action of sufentanil (0.5 microM), morphine (5 microM), and DAME (5 microM), but not U-50,488H (10 microM). The action of U-50,488H was partially blocked by the antagonist naltrexone (25 microM). Stereo-selectivity of agonist action was confirmed by the failure of dextrorphan (50 microM), an inactive opioid isomer, to inhibit the release of substance P.(ABSTRACT TRUNCATED AT 250 WORDS)

Regional serotonin and dopamine activity : sensitivity to amphetamine and aggressive behavior in mice

We were interested in comparing the effect of distinctive attack and defeat experience on regional monoamine turnover and on d-amphetamine (AMPH) sensitivity. Male mice were randomly assigned to groups that received zero, one, or ten defeat experiences or that attacked an intruder zero, one, or ten times. Twenty-four hours after the last attack or defeat, mice were administered one of four AMPH doses (0, 0.1, 1.0, or 10.0 mg/kg i.p.) and tested in a resident-intruder confrontation. A separate group of mice were sacrificed immediately after the last attack or defeat. Using the high-performance liquid chromatography (HPLC) technique, dihydroxyphenylacetic acid (DOPAC):dopamine (DA) and 5-hydroxyindolacetic acid (5HIAA):serotonin (5HT) were measured in micropunched sections of the corpus striatum, amygdala, and nucleus accumbens. A single defeat decreased DA turnover in the amygdala, whereas one attack experience increased DA and 5HT turnover in the n. accumbens and amygdala, respectively. AMPH sensitivity was enhanced following acute defeat. In animals that had repeated agonistic experience, acute attack or defeat did not alter DOPAC:DA or 5HIAA:5HT ratios. Mice with attack experience were more sensitive to AMPH following multiple rather than a single aggressive encounter. The distinct effects of defeat and attack experience on mesolimbic DA activity and on AMPH sensitivity suggest that monaminergic fibers terminating in the n. accumbens can be functionally dissociated from fibers terminating in the amygdala.

Social defeat stress increases expression of μ-opioid receptor mRNA in rat ventral tegmental area

Prompted by previous studies linking social defeat stress to changes in opioid antinociception, we evaluated the expression of mu-opioid receptor (MOR)-encoding mRNA in selected rat brain areas as a function of this type of stress. Because opioids mediate significant regulatory activities of brain dopamine neurons, dopaminergic loci in the ventral tegmental area (VTA) and substantia nigra (SN) were selected for analysis. Within 30 min after social defeat stress, the level of MOR-encoding mRNA, as detected and quantified by in situ hybridization histochemistry, increased in the lateral VTA and this increase was present for at least 6 h. In contrast, defeat stress was without effect on the expression of MOR-encoding mRNA in the SN. These data suggest that stress-induced alteration of MOR-encoding mRNA expression in the VTA may be involved in the consequences of social defeat stress.

Behavioral sensitization to cocaine after a brief social stress is accompanied by changes in fos expression in the murine brainstem.

The objective of the present study was to determine how c-fos gene expression in brainstem structures after a brief episode of social defeat stress is related to behavioral sensitization to cocaine challenge. Social stress was defined as defeat in a brief confrontation with an aggressive resident mouse and subsequent 20-min exposure to the residents threats behind a protective screen. Mice were treated with cocaine (40 mg/kg, i.p.) immediately or 1 week after social defeat stress. Fos-like immunoreactive (Fos-LI) cell nuclei were analyzed in the ventral tegmental area (VTA), dorsal raphe nucleus (DR), periaqueductal grey area (PAG) and locus coeruleus (LC). One episode of social stress induced behavioral sensitization to cocaine as indicated by an augmented locomotor response to a challenge injection 7 days after a single defeat. In naive mice, social stress markedly increased the number of Fos-LI nuclei in the DR, PAG and LC, but not in the VTA. Similarly, cocaine administration resulted in a significantly increased number of Fos-LI nuclei in the same areas. Administration of cocaine immediately following social defeat significantly reduced the number of Fos-LI nuclei in the DR, PAG and LC. Cocaine-induced Fos expression returned in the PAG and DR, but not in the LC, 1 week after social stress. In conclusion, the present results suggest that the presence of brainstem Fos be related to the ability to express stress-induced behavioral sensitization to cocaine.

Spinal dynorphin involvement in the analgesia of pregnancy: effects of intrathecal dynorphin antisera.

In both rats and humans there is an analgesia associated with pregnancy. This analgesia is spinally mediated and involves the kappa type of opiate receptor. The current study demonstrates that intrathecal administration of high affinity dynorphin antibodies produces a significant reduction in jump thresholds during pregnancy (day 20). The administration of pre-adsorbed antisera fails to produce this effect. These results support the hypothesis that a spinal dynorphin/kappa opiate receptor system is activated during gestation.

Distribution of tachykinin- and opioid-expressing neurons in the hamster solitary nucleus: an immuno- and in situ hybridization histochemical study

In several sensory systems, tachykinin- and opioid-expressing neurons functionally interact and influence the processing of afferent information. To determine whether a similar relationship exists for the processing of general and special (gustatory) visceral afferent information, the present study mapped the distributions of these two neuronal phenotypes within the nucleus of the solitary tract (NST) of the hamster by employing a combination of immuno- and in situ hybridization histochemistry (ISHH). The hamster was chosen because it is frequently used as a model in taste studies, yet there is a relative dearth of data about peptide expression or the classical neurotransmitters in the brainstem of this animal. The immunohistochemical analyses employed 2 highly selective antisera directed towards the prototypical tachykinin and opioid peptides, i.e. substance P (SP) and methionine enkephalin (ENK), respectively. Intense staining of fibers and preterminal/terminal puncta was concentrated in the rostral pole or gustatory zone of the NST. SP-, but not ENK-like immunoreactivity was also observed in long courses of axon bundles traversing the brainstem enroute to the NST. Local application of colchicine engendered the appearance of a moderate number of SP-positive somata that were mostly clustered in the medial, central and intermediate subnuclei, as well as being scattered throughout the remainder of the NST, including the gustatory zone. A low number of isolated ENK-positive somata were also observed throughout the NST. The somal areas of the SP- and ENK-positive somata averaged 86.3 and 81.8 microns 2, respectively. The ISHH studies were performed using 2 selective oligodeoxynucleotide probes with complementary sequences to mRNAs encoding gamma-preprotachykinin (PPT) and preproenkephalin (PPE) molecules. Overall, the cellular expression of PPT mRNA within the NST corresponded both in distribution and in number to those identified by immunohistochemical analyses using anti-SP serum. In contrast, ISHH analyses monitored a significantly greater number of PPE-expressing somata in the medial, central, intermediate and ventrolateral nuclei than were ENK immunoreactive. These findings indicate that tachykinin and opioid peptide phenotypes are represented in neurons throughout the hamster NST and suggest a functional role for PPT- and PPE-related peptide forms in the modulation of afferent general visceral and gustatory information.