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How obesity exacerbates migraine and other pain disorders remains unknown. Trigeminal nociceptive processing, crucial in migraine pathophysiology, is abnormal in mice with diet induced obesity. However, it is not known if this is also true in genetic models of obesity. We hypothesized that obese mice, regardless of the model, have trigeminal hyperalgesia. To test this, we first evaluated trigeminal thermal nociception in leptin deficient (ob/ob) and control mice using an operant thermal assay. Unexpectedly, we found significant hypoalgesia in ob/ob mice. Because thermal hypoalgesia also occurs in mice lacking the transient receptor potential vanilloid 1 channel (TRPV1), we tested capsaicin-evoked trigeminal nociception. Ob/ob and control mice had similar capsaicin-evoked nocifensive behaviors, but ob/ob mice were significantly less active after a facial injection of capsaicin than were diet-induced obese mice or lean controls. Conditioned place aversion in response to trigeminal stimulation with capsaicin was similar in both genotypes, indicating normal negative affect and pain avoidance. Supporting this, we found no difference in TRPV1 expression in the trigeminal ganglia of ob/ob and control mice. Finally, we assessed the possible contribution of hyperphagia, a hallmark of leptin deficiency, to the behavior observed in the operant assay. Ob/ob and lean control mice had similar reduction of intake when quinine or capsaicin was added to the sweetened milk, excluding a significant contribution of hyperphagia. In summary, ob/ob mice, unlike mice with diet-induced obesity, have trigeminal thermal hypoalgesia but normal responses to capsaicin, suggesting specificity in the mechanisms by which leptin acts in pain processing.

The broad use and misuse of prescription opioids during pregnancy has resulted in a surge of infants diagnosed with Neonatal Opioid Withdrawal Syndrome (NOWS). Short-term irritability and neurological complications are hallmarks of NOWS, but the long-term consequences are unknown. Our newly-developed preclinical model of oxycodone self-administration enables adult female rats to readily drink oxycodone (0.06-0.12 mg/ml, ∼10/mg/kg/day) continuously before and during pregnancy and after delivery, to achieve similar liquid intake in oxycodone moms relative to water-only controls. Oxycodone levels were detected in the serum of mothers and pups. Growth parameters in dams and pups, and litter mass and size were similar to controls. Maternal behavior at postnatal day 1 (PN1) was unchanged by perinatal oxycodone consumption. Regarding the plantar thermal response, there were no differences in paw retraction latency between oxycodone and control pups at PN2 or PN14. Oxycodone and control pups had similar motor coordination, cliff avoidance, righting time, pivoting, and olfactory spatial learning from PN3 through PN13. Separation-induced ultrasonic vocalizations at PN8 revealed higher call frequency in oxycodone pups relative to controls. Finally, during naltrexone precipitated withdrawal at PN9, oxycodone males vocalized more than control pups, consistent with a previously-published withdrawal phenotype. Thus, our rat model of continuous oral oxycodone self-administration in pregnancy shows exacerbated affect/social communication in pups in a sex-dependent manner but spared cognition and locomotion. Our preclinical, high face validity NOWS model reproduces key aspects of human opioid use during pregnancy, enabling longitudinal analysis of how maternal oxycodone changes emotional behavior in the offspring.HIGHLIGHTSFemale rats self-administered oxycodone at clinically relevant doses before and during pregnancy and for the first two weeks after parturition.Both dams and pups, for the14 day postnatal experimental period, had detectable levels of oxycodone in their bloodDams drinking oxycodone only or water only did not differ in weight gain, water intake, or the number of pups born and their pups did not differ in weight throughout.Sensory and motor function in the pups was not altered, nor was hippocampal dependent spatial learning.Oxycodone exposed pups were physically dependent and displayed increased withdrawal behaviors with or without the opioid antagonist naltrexone.Pups expressed more negative affect, expressed by increased ultrasonic vocalizations, following naltrexone precipitated withdrawal or when separated from their mother.

The selective serotonin reuptake inhibitor fluoxetine and the 5-HT1A receptor agonist buspirone are used to treat depression and anxiety. Previously we demonstrated that chronic stress during pregnancy (prenatal stress) in rats, used as a model of maternal depression risk, increased inflammatory pain and depressive-like behavior in the offspring; buspirone injected to pregnant dams was protective. Clinically, the addition of buspirone to fluoxetine increases the latter’s efficacy in treating depression in patients. Here, we investigated the influence of repeated prenatal injections of fluoxetine, buspirone or their combination on pain- and depressive-like behaviors in prenatally stressed young male and female rats. Prenatal stress augmented depressive-like behavior and both thermal and inflammatory pain (formalin test), replicating our prior findings, and increased basal levels of corticosterone in the blood plasma. Both drugs and their combination reduced the effects of prenatal stress on thermal pain and depressive-like behavior independently of sex. The combination of fluoxetine and buspirone, compared with fluoxetine, was more antinociceptive in the hot plate test in both sexes, and when compared with buspirone, was more antinociceptive only in males. A detailed study of the time-course of formalin-induced pain showed a nuanced effect of these drugs that was sex-dependent. The combination of the two drugs was less effective in females than males during the initial acute phase of nociceptive behavior in flexing + shaking behaviors, whereas that combination was more effective than fluoxetine alone in the first acute phase of licking behavior in females. The antinociceptive effect of buspirone dominated that of the drug combination and of fluoxetine alone, especially during the interphase of the formalin test in both sexes for both flexing + shaking and licking, suggesting a more effective prenatal action of buspirone on the development of a descending serotonergic inhibitory system modulating pain in the spinal cord dorsal horn neurons. Our results indicate that inflammatory pain-like responses integrated at the spinal level in males were more vulnerable to prenatal stress than females. In licking, the antinociceptive effect of fluoxetine and drug combination in the interphase was more in males than females. The data underscore the importance of considering sexual dimorphism when using drug therapy.

The effects of prenatal stress on the immunocytochemical reaction for serotonin (5-HT) in the dorsal raphe nucleus (DRN) were studied in seven-day-old male rat pups subjected to pain in the formalin test (inflammatory pain; controls received injections of physiological saline. Prenatal stress enhanced the animals’ pain behavior in the formalin test. Prenatal stress decreased the numbers of 5-HT-immunoreactive cells as compared to those in prenatally unstressed animals, both in rat pups with inflammatory pain and in controls. The formalin treatment did not alter the number of 5-HT-immunoreactive cells in the DRN in rat pups in either prenatally stressed or unstressed animals. Thus, we demonstrate that 5-HT-ergic neurons in the brainstem DRN in seven-day-old rat pups are the targets of prenatal stress. We suggest that stress due to separation from the mother during the experiment masked differences in the immunocytochemical parameters in rat pups with inflammatory pain and in controls.

In the adult, immune and neural processes jointly modulate pain. During development, both are in transition and little is known about the role that the immune system plays in pain processing in infants and children. The objective of this study was to determine if inhibition or augmentation of the immune system would alter pain processing in the infant rat, as it does in the adult. In Experiment 1, rat pups aged 3, 10, or 21 (PN3, PN10, and PN21) days of age were pretreated with NS398 (selective cyclooxygenase (COX)-2 inhibitor) or SC560 (selective COX-1 inhibitor) and tested in the intraplantar formalin test to assess effects of COX inhibition on nociception. Neither drug had an effect on the behavioral response at PN3 or PN10 pups but both drugs attenuated nociceptive scores in PN21 pups. cFos expression in the spinal cord likewise was reduced only at PN21. In Experiment 2, pups were injected with lipopolysaccharide (LPS) prior to the formalin test at PN3 or PN21. LPS increased the nociceptive response more robustly at PN21 than at PN3, while increasing cytokine mRNA equally at both ages. The augmentation of pain responding at PN21 was largely during the late stages of the formalin test, as reported in the adult. These data support previous findings demonstrating late maturing immune modulation of nociceptive behaviors.

How exogenous estrogen affects inflammatory responses is poorly understood despite the large numbers of women receiving estrogen-alone hormone therapy. The aim of this study was to determine if estradiol alters injury- or inflammation-induced nociceptive responses after carrageenan administration in females and whether its effects are mediated through cyclo-oxygenase (COX) and prostaglandins (PG). To this end, paw withdrawal latencies and serum levels of PGE2 and PGD2 were measured in rats treated with estradiol (0, 10, 20, and 30%) and/or SC560 (COX-1 inhibitor) or NS398 (COX-2 inhibitor) after intraplantar carrageenan administration. Estradiol significantly increased withdrawal latencies before (baseline condition) and after carrageenan administration to one hindpaw. NS398 was anti-nociceptive only in carrageenan treated animals. SC560 increased withdrawal latencies in both paws at 1 and 5hours after carrageenan administration. Co-administration of estradiol and NS398, but not SC560, was additive except for a prolonged anti-nociceptive effects of estradiol combined with NS398. The anti-nociceptive effect extended beyond that observed with either drug or estradiol alone at the 5-hour time point. Estradiol had no significant effect on PGE2 serum levels, but both COX antagonists decreased them. Although neither estradiol nor the COX inhibitors alone had an effect on PGD2 serum levels, co-administration of NS398 and estradiol significantly elevated PGD2 levels. Taken together, our results suggest that estradiol is anti-nociceptive in the thermal test and reduces carrageenan-induced hyperalgesia. These effects are minimally altered through PG-mediated mechanisms.

Estrogen modulates pain perception but how it does so is not fully understood. The aim of this study was to determine if estradiol reduces nociceptive responses in part via hypothalamic-pituitary-adrenal (HPA) axis regulation of cyclooxygenase (COX)-1/COX-2 activity. The first study examined the effects of estradiol (20%) or vehicle with concurrent injection nonsteroidal antiinflammatory drugs (NSAIDs) on formalin-induced nociceptive responding (flinching) in ovariectomized (OVX) rats. The drugs were ibuprofen (COX-1 and COX-2 inhibitor), SC560 (COX-1 inhibitor), or NS398 (COX-2 inhibitor). In a second study, estradiol's effects on formalin-induced nociception were tested in adrenalectomized (ADX), OVX, and ADX+OVX rats. Serum levels of prostaglandins (PG) PGE(2) and corticosterone were measured. Estradiol significantly decreased nociceptive responses in OVX rats with effects during both the first and the second phase of the formalin test. The nonsteroidal antiinflammatory drugs (NSAIDs) did not alter nociception at the doses used here. Adrenalectomy neither altered flinching responses in female rats nor reversed estradiol-induced antinociceptive responses. Estradiol alone had no effect on corticosterone (CORT) or prostaglandin levels after the formalin test, dissociating the effects of estradiol on behavior and these serum markers. Ibuprofen and NS398 significantly reduced PGE2 levels. CORT was not decreased by OVX surgery or by estradiol below that of ADX. Only IBU significantly increased corticosterone levels. Taken together, our results suggest that estradiol-induced antinociception in female rats is independent of COX activity and HPA axis activation.

Human infants are often exposed to opiates chronically but the mechanisms by which opiates induce dependence in the infant are not well studied. In the adult the brain regions involved in the physical signs of opiate withdrawal include the periaqueductal gray area, the locus coeruleus, amygdala, ventral tegmental area, nucleus accumbens, hypothalamus, and spinal cord. Microinjection studies show that many of these brain regions are involved in opiate withdrawal in the infant rat. Our goal here was to determine if these regions become metabolically active during physical withdrawal from morphine in the infant rat as they do in the adult. Following chronic morphine or saline treatment, withdrawal was precipitated in 7-day-old pups with the opiate antagonist naltrexone. Cells positive for Fos-like immunoreactivity were quantified within select brain regions. Increased Fos-like labeled cells were found in the periaqueductal gray, nucleus accumbens, locus coeruleus, and spinal cord. These are consistent with other studies showing that the neural circuits underlying the physical signs of opiate withdrawal are similar in the infant and adult.

Neonates respond to noxious stimuli at or before birth, but the organization of nociceptive systems changes well into postnatal life. It is unknown how nociceptive information is processed in the immature animal and, specifically, whether noxious stimulation is transmitted by glutamatergic circuits, known to play an important role in nociception in the adult. Both N-methyl-d-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors are found within the neonatal spinal cord, but in immature form, and when they become involved in pain processing in vivo is not known. The objective was to determine the age-related changes in the involvement of spinal NMDA and AMPA receptors in formalin-induced nociception during early life. Because the formalin test provides a measure of immediate nociceptive responding (first phase) and of peripheral and central sensitization (second phase), a second aim was to determine if there is specificity of the effects to either phase. NMDA antagonists (MK801, AP5) or an AMPA antagonist (YM872) was administered intrathecally, and pups were assessed in the formalin test behaviorally and by Fos expression within the spinal cords of 3-, 10-, and 21-day-old rats. The NMDA antagonists attenuated formalin-induced behavioral responses at the youngest age tested with some selectivity for the second phase of responding. MK-801 did not induce motor impairment at any age. YM872 also attenuated formalin-induced nociceptive responses at all ages throughout the test session, although there was some motor impairment in the 3-day-old subjects. Spinal administration of either YM872 or MK-801 reduced Fos expression in the spinal cord at all ages. These data suggest that spinal NMDA and AMPA receptor are functional and involved in formalin-induced nociception throughout development.

When pregnant dams are stressed, there is a resultant alteration in brain development and behavior in their offspring. Prior work has shown increased nociceptive responses in adolescent or adult rats born of stressed dams. However, the age at which those changes first occur is not known. The aim of the present study was to evaluate the effect of prenatal stress on pain sensitivity in the formalin test in 7-day-old rats, behaviorally and by fos-like immunoreactivity (Fos-LI) in the lumbar spinal cord dorsal horn. The behavioral response to intraplantar injection of formalin is represented by two nociceptive phases separated by an interphase during which nociceptive responses decrease; the interphase is not seen until the start of the third postnatal week and appears as descending inhibitory monoaminergic systems develop. Prenatally stressed infants showed increased nociceptive responses in the second, tonic phase and a large increase in the number of formalin-induced Fos-LI neurons in the lumbar dorsal horn, a result consistent with the behavioral data. The increased nociception in prenatally stressed 7-day-old pups may be associated with the decrease in the intensity of serotonin-like immunoreactivity and density of serotonergic cells in the lumbar spinal cord dorsal horn and the dorsal raphe nucleus reported earlier.

Young animals respond to threatening stimuli in an age-specific way. Their endocrine and behavioral responses reflect the potential threat of the situation at a given age. The aim of the present study was to determine whether corticotropin-releasing factor (CRF) is involved in the endocrine and behavioral responses to threat and their developmental changes in young rats. Preweaning 14-day-old and postweaning 26-day-old rats were exposed to two age-specific threats, cat odor and an adult male rat. The acute behavioral response was determined during exposure. After exposure, the time courses of the corticosterone response and of CRF expression in the paraventricular nucleus of the hypothalamus (PVN) and in extrahypothalamic areas were assessed. Preweaning rats became immobile when exposed to cat odor or the male rat, whereas postweaning rats became immobile to cat odor only. Male exposure increased serum corticosterone levels in 14-day-old rats, but cat odor failed to increase levels at either age. Exposure induced elevation of CRF mRNA levels in the PVN that paralleled changes in corticosterone levels. CRF may thus play a role in endocrine regulation and its developmental changes during early life. Neither cat odor nor the adult male altered CRF mRNA levels in the bed nucleus of the stria terminalis (BNST) or the amygdala, but both stimuli increased levels in the hippocampus. Hippocampal CRF mRNA expression levels did not parallel cat odor or male-induced immobility, indicating that CRF is not involved in this response in young rats but may be involved in aspects of learning and memory.

It is unclear how neonates respond to noxious stimuli. This study examined the role of neurokinin peptides in 3- and 21-day-old rat pups using the preprotachykinin-A (PPTA) knockout mouse, lacking neurokinin A and substance P. We assessed pain behaviors of these mice before the neurokinin system is putatively active, 3 days after birth, and after this system is active, 21 days after birth. The lack of these peptides failed to alter behavioral responses to nociceptive stimulation in the 3-day-old mice. The 21-day-old mice lacking these peptides were less responsive to 5 microl 2% formalin and to high intensity thermal and mechanical stimuli. Thus, the neurokinins appear not to be an important mechanism in the processing of nociceptive information in the infant.

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.

Opiate withdrawal behaviors in the infant differ from those of the adult. The neural circuitry underlying opioid withdrawal in the adult rat is well defined and includes the locus coeruleus (LC) and periaqueductal gray (PAG), with a minor role of the amygdala. Because the different behaviors that constitute the infant syndrome may be mediated by different neural circuits, we tested the hypothesis that these three sites are involved in opiate withdrawal. Pups were injected with morphine from day 1- 6 after birth (b.i.d.) and on the morning of the seventh day. Withdrawal was then elicited by local injection of the opioid antagonist methylnaloxonium into the LC, PAG, or amyg- dala. Withdrawal signs were precipitated in a dose-dependent manner following injec- tion into the LC or PAG, but not the amygdala. The withdrawal behaviors elicited from the LC and PAG included both the same and different behaviors. The results support the hypothesis that the neural circuitry mediating opiate withdrawal behaviors is similar in infant and adult animals, but the behaviors expressed are age-specific. Synapse 39: 139 -151, 2001. © 2001 Wiley-Liss, Inc.

Offspring of women exposed to opiate drugs such as heroin and methadone during pregnancy have a high incidence of morbidity and mortality. Infants also show opiate withdrawal. In this study, we examined the behavioral effects of precipitated withdrawal in morphine-dependent fetal rats at gestational day (GD) 20. The dam was implanted on GD 14 with a pellet containing 75.0 mg of morphine. On GD 20, the dam underwent chemoyelotomy at L1/L2. The uterine horns were externalized and four subject fetuses were selected for behavioral observation, two from each uterine horn. The fetus was then injected subcutaneously with either saline or naltrexone (1.0 mg/kg) and the behaviors of the fetus recorded every 15 sec for 20 min. The results show that naltrexone injected fetuses that had been chronically exposed to morphine demonstrated increased limb and mouth movements, face wiping, and body curls, and spent less time quiet as compared with control fetuses. These results indicate that a morphine withdrawal-like syndrome occurs in the fetal rat.

Endomorphin-1 and endomorphin-2 are recently described peptides with high affinity and specificity for the mu opioid receptor. They are believed to be the endogenous ligands for that receptor. We describe the maturation of the endomorphins in brain and spinal cord using recently characterized antibodies to each. Endomorphin-1-like immunoreactivity was examined in brain, focusing on the periaqueductal gray of the midbrain and the diagonal band of Broca; endomorphin-2-like immunoreactivity is reported for the medulla and spinal cord. In these regions, and in all other regions studied but not described in this paper, the endomorphins were not seen at birth or at 3 days of age. Staining was present in 7-day-old and older animals. At these early ages, the pattern and density of staining are not fully developed, but appear complete by 21 days of age. The results suggest that both endomorphin-1 and endomorphin-2 develop relatively late compared to other opioid peptides.

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.

Recent evidence has shown that infant rats undergo precipitated withdrawal following chronic postnatal injection of morphine. In this study we examined whether or not infants exposed to methadone prenatally via the placental blood supply and postnatally via the dam’s milk would also experience precipitated withdrawal. Dam’s were implanted on gestational day 14 with osmotic minipumps containing one of two concentrations of methadone to supply the opiate throughout gestation and the first postnatal week. Nontreated and pair-fed controls were used. On postnatal day 7, pups were injected with naltrexone and their locomotor activity and ultrasonic vocalizations measured. Methadone exposed pups were more active and vocalized more when injected with naltrexone than with saline. The controls did not show these behavioral changes. The milk of methadone-exposed dams apparently contains sufficient quantities of the opiate for dependence to develop. The results are consistent with other data that demonstrate that very young rat pups can experience an opiate abstinence syndrome that includes increased behavioral activation.

The dorsal midbrain including the periaqueductal gray (PAG) is involved in the control of threat-induced vocalizations and other behavioral and autonomic defensive responses in adult animals. Little is known of its function early in life. The present study examined the ability of kainate receptor stimulation in these midbrain areas to trigger behavioral and physiological responses during the first three postnatal weeks in the rat. Kainate (0.03-0.3 nmol) was infused into the dorsal midbrain of postnatal day 7 (P7), P14 and P21 rat pups. At P7, subjects exhibited only a brief period of locomotor activation immediately following infusion of kainate. There were no changes in the heart rate or in any other behavioral measures, including their production of ultrasonic vocalizations. At P14, kainate induced adult-typical escape behaviors consisting of running and jumping, increases in the duration of time spent immobile, and increases in heart rate. At P21, subjects given kainate exhibited escape behavior coupled with elevated heart rate and immobility coupled with decreased heart rate. P14 and P21 subjects produced only small, non-significant increases in their production of ultrasonic vocalizations. These results indicate that kainate receptor stimulation in the dorsal midbrain does not mediate most adult-typical, threat-induced responses until sometime during the second postnatal week in the rat.

This study examined the normal development of neuronal activity in the suprachiasmatic nuclei (SCN) of rats between age 3-60 days, using Fos protein-like immunoreactivity (Fos-LI) as a marker. At age 3 days, Fos-positive nuclei are sparsely distributed throughout the SCN. Between age 3-10 days, the density of labeled nuclei increases significantly. Fos-LI labeling is maximal at 10 days. Between age 10-14 days, the number of labeled nuclei decreases and remains relatively constant thereafter, although the intensity of the reaction product diminishes as the animal matures. By age 60 days, the number of Fos-LI labeled nuclei in the SCN is substantially decreased and is essentially the same as in the 3-day-old rat. The appearance of Fos-LI nuclei in the SCN during development appears to reflect the development of visual system afferents to the nucleus as well as the development of intrinsic SCN synaptology.

The present study examined the maturation of nociceptive primary afferents using expression of Fos-like immunoreactivity in the second-order spinal cord neurons as an anatomical and functional marker. Pinch, immersion in hot water, or formalin injection applied to the hindpaw was used as the peripheral noxious stimulus in awake 0-, 1-, 2-, 3-, and 14-day-old rat pups. On the day of birth, all 3 stimuli elicited expression of the Fos protein in dorsal horn cells indicating that nociceptive primary afferents are functional at this age. The expression of the Fos protein was related to the intensity of stimulation since greater injection volumes of formalin or prolonged application of the thermal stimulus increased the number of stained nuclei. The number of stained nuclei was age dependent and older pups exhibited a greater number of stained nuclei. The results of this study are consistent with electrophysiological studies that have demonstrated that the primary nociceptive afferents continue to mature during the rats' postnatal life. Furthermore, the number of Fos immunoreactive neurons in immature rats are age and stimulus-intensity dependent.

mu Opioids have been shown to produce analgesia and to be reinforcing during the first week of life in the rat. kappa Opioids also have analgesic actions in both the infant and adult, but can be aversive in the mature animal. We examined the aversive effects of the kappa opioid agonist U50,488 during the first postnatal week in the rat pup in three ways. In the first experiment, U50,488, injected peripherally (1.0-30.0 mg/kg), was paired with an odor and pups were tested 8 h later for positional preference for avoidance of that odor. This task is similar to conditioned preference/aversion tests used with adult animals. Both 3- and 7-day-old pups learned to avoid the odor adulterated side at the two higher doses. When exposed to odors previously associated with U50,488, pups at both ages decreased locomotor activity. In a second experiment, acute treatment with U50,488 increased ultrasonic distress vocalizations (USV) equally at 3 and 7 days of age, increased locomotor activity, and decreased rectal temperature. Neither of the latter two effects was correlated with the increase in USV production. The third experiment showed that conditioned odor cues increased USV 8 h later in 3- and 7-day-old pups at 1.0-10.0 mg/kg without changes in activity or rectal temperature. The results from these studies suggest that U50,488 can produce aversions in the neonatal rat pup as it does in the adult.

In the fetal, infant, and adult rat, injury induces a well-defined behavioral response and induces c-fos expression in the spinal cord dorsal horn. There is more limited information about the processing of noxious stimulation in the infant brain. We describe here the appearance of the Fos protein in the brain of fetal and infant rats following formalin-induced injury. Regions were chosen for analysis with a special focus on brain loci that express c-fos in the adult. No Fos positive cells were found in the brains of fetuses; newborns did not show increased Fos expression after formalin injection in any structure examined. At 3 and 14 days of age, there was a significant increase in Fos staining induced by formalin in the ventral lateral medulla. In contrast, paraventricular and medial dorsal nuclei of the thalamus, the paraventricular nucleus of the hypothalamus, and periaqueductal gray of the midbrain showed increased levels of Fos protein only at 14 days of age. We hypothesize that this developmental pattern is related not only to the maturation of pain perception but also to development of autonomic and defensive reactions to pain in the infant. Perspective Because the infant processes pain differently than the adult, knowledge of those differences informs pediatric clinical practice. Using Fos expression as a marker of neural activity in the rat, we show that the pattern of brain activation is immature at birth but is in place by 14 days of age.

To assess the development of the reinforcing properties of cocaine, 3- to 4-day-old rat pups were administered cocaine s.c (3.0-30.0 mg/kg), intracerebroventricularly (3.0-10.0 micrograms/injection), or directly to the nucleus accumbens (1-25 micrograms/injection) in the presence of a distinctive odor. Six to 8 h later, pups were allowed a choice between the paired odor and a non-paired odor. Compared to saline treated controls, pups demonstrated a preference for the odor paired with cocaine for each route of administration. Taken together with previous work demonstrating that cocaine increases responding for electrical self-stimulation of the basal forebrain, these results suggest that cocaine is rewarding in the infant animal and that mesolimbic structures may mediate, at least in part, those reinforcing properties.

The present study was undertaken to assess the functional ontogeny of alpha 2-noradrenergic receptors in the hypothalamic paraventricular nucleus (PVN) that mediate noradrenergic stimulation of feeding in the rat. Rat pups, ranging in age from 2 to 15 days, were removed from their mothers and implanted with a brain cannula directed unilaterally at the PVN or third ventricle. On the following day, each pup was implanted with an intra-oral cannula for oral infusion of milk or water that could be swallowed or rejected. Following a 1-h period of satiation, each pup received an intracerebral injection of saline, or a single dose of norepinephrine (NE, 0.01-100.0 nmol) or the alpha 2-noradrenergic receptor agonist clonidine (0.01-1.0 nmol). Milk or water intake was then assessed following a 1-h period of infusion. Injection of NE into the PVN and third ventricle significantly enhanced milk intake at 2 days of age. NE was 10-fold more potent in the PVN than in the ventricle. While paraventricular injections of NE stimulated milk and water intake equally at 2 days of age, NE produced a greater stimulation of milk than water intake at 15 days of age. Like NE, clonidine significantly enhanced milk intake at 2 days of age following injection into the PVN. Collectively, these findings suggest that alpha 2-noradrenergic receptors, mediating noradrenergic stimulation of feeding, are functionally mature very early in the postnatal development of the rat. Moreover, consistent with evidence in the adult rat, these findings indicate that alpha 2-noradrenergic receptors relevant to feeding are located in the vicinity of the PVN.

This study examined cellular activity in the hypothalamus of developing rats in response to cold environmental temperatures. The appearance of the nuclear protein, Fos, in response to cold ambient temperatures in rats three to 35 days of age was used as a marker of neuronal activation. Fos-positive nuclei were first seen in response to cold ambient temperatures in the ventromedial nucleus at three days of age, the paraventricular nucleus at eight days of age, the preoptic-anterior hypothalamus at 10 days of age and the anterior hypothalamic nucleus at 21 days of age. The rectal temperature of the 10-day-old pups dropped by less than half that measured in the nine-day-old pups after 1 h in the cold. It is possible that the activation of neurons in the preoptic-anterior hypothalamus in the 10-day-old animal may contribute to the decreased hypothermia observed in the 10-day-old after 1 h in the cold. The high density of Fos-like immunoreactive-labeled nuclei in the preoptic-anterior hypothalamus in the 10-day-old rats that were exposed to the cold environment indicates increased neuronal activity at this site. The results suggest that, at least in the preoptic-anterior hypothalamus, the appearance of the cellular response to cold ambient temperatures is coincidental with a thermogenic response.

When given peripherally, morphine produces a conditioned place preference (CPP) in adult and infant rats. In adults, morphine injected into the ventral tegmental area (VTA) is reinforcing and likely acts by activating mesolimbic dopamine neurons. Little is known of the neurobiological basis of reinforcement in the immature animal. To test whether the neural substrate of drug reinforcement is similar in adults and infants, 4-day-old pups were injected with one of 4 doses (0.05, 0.15, 0.45, 1.35 micrograms) of morphine or the vehicle directly into the VTA and were immediately confined to an odor-cued environment for 30 min. They were then tested for a preference between the cued environment and unadulterated wood shavings. The low dose of morphine (0.05 micrograms) injected into the VTA significantly increased duration of time spent in the conditioned environment, demonstrating conditioned preference for the conditioned area over the unscented area; the most effective injection sites were directly into the VTA. Stimulation of structures just outside the VTA, and higher doses of morphine were ineffective. On the basis of these findings we conclude that the neural substrates of reward in the neonate are similar to those of the adult.

The possibility was investigated that specific opioid receptor types might selectively alter the production of isolation-induced ultrasonic vocalizations. Intracisternal injections of mu, delta and kappa opioid receptor agonists were administered to isolated 10-day-old rat pups. The mu receptor agonist [D-Ala2-NMe-Phe4-Gly-ol]-enkephalin (DAMGO) and delta receptor agonist [D-Pen2, D-Pen5]-enkephalin (DPDPE) both reduced the rate of isolation-induced ultrasonic calling in the absence of sedation. The kappa receptor agonist U50,488 had the opposite effect, significantly raising the rate of vocalization. Fourteen-day-old pups, with a larger delta receptor population, showed a greater sensitivity to DPDPE than was seen in the younger animals.

Acute administration of naloxone to preweanling rats does not attenuate independent ingestion of milk until 14 days of age suggesting that the full expression of an endogenous opioid system(s), regulating feeding rats, is not complete prior to this age. The present study was undertaken to examine the functional ontogeny of opioid receptors mediating opiate-induced feeding in rats. Rat pups, satiated with milk, were given intraperitoneal injections of the opiate receptor agonist, morphine, and were allowed free access to milk. Morphine stimulated the intake of milk at 3, 5, 7, 14 and 21 days of age, within 2 hr of injection. A time-course analysis in 7-day-old pups showed greater enhancement of intake between hours 2 and 4, than between hours 0 and 2, for large doses of morphine (0.3 and 1.0 mg kg ) suggesting that morphine-induced behavioral depression, which was observed early in the test session, confounded intake at earlier hours. Administration of the opiate receptor antagonist, naltrexone, produced no effect on intake of its own, but blocked the stimulation of intake by morphine in 5-day-old pups confirming that the effect of morphine on the intake of milk was mediated by opioid receptors. Thus, while a functional endogenous opioid system(s), regulating feeding in rats, is not fully mature until 14 days postpartum, the present results suggest that opioid receptors mediating feeding are functional very early in the postnatal development of the rat.

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 estradiol's analgesic effect in Phase II, however, estradiol did not reverse progesterone's 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 estradiol's 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.

Kappa opioid receptors stimulation with U50,488 is known to modulate behaviors during the early postnatal period, but the specific neuroanatomical locus of many of these effects is unexplored. In the present study, we infused U50,488 into the midbrain periaqueductal gray (PAG) and investigated the effects of this drug on behavior and heart rate of 1-, 2-, and 3-week-old rats. U50,488 increased activity most potently in 1- and 2-week-old subjects. Ultrasonic vocalization (USV) production was increased in 1-week-old subjects, but not in 2- or 3-week-old pups. Heart rate changes were similarly seen in younger aged subjects. At 1 week, U50,488 decreased heart rate, but at 2 weeks it increased heart rate. There was no effect of this drug on heart rate at 3 weeks. At 1 week, USVs were more potently elicited from dorsal than lateral PAG infusion sites. No other site-specific effects within the PAG were seen. The age-related decline in behavioral effects elicited by U50,488 is consistent with other published reports, and to the extent that kappa receptor activity mediates infant separation responses, implicates the PAG as a modulator of those responses.

Exposure to a deadly threat, an adult male rat, induced the release of corticosterone in 14-day-old rat pups. The endocrine stress response was decreased when the pups were reunited with their mother immediately after exposure. These findings demonstrate that social variables can reduce the consequences of an aversive experience.

N-methyl-D-aspartate (NMDA) receptor antagonists are effective in inhibiting the development of morphine tolerance in adult rats. But NMDA receptors undergo dramatic change during the first few weeks of the postnatal life in the rat, and it is unknown whether NMDA receptor antagonists can inhibit the acquisition of opiate tolerance in the developing organism. Here, we investigated the effects of two NMDA receptor antagonists MK-801 and dextromethorphan on the development of morphine tolerance in 7-, 14-, and 21-day-old rats. NMDA receptor antagonists are not effective in attenuating morphine tolerance in the neonatal rat whereas they were partially effected in the 14-day-old and fully effective in rats as old or older than 21 days of age. These data suggest that there exists a transition age, around the second postnatal week in the rat, for the NMDA receptor to play a role in the development of morphine tolerance.

Stress activates endogenous opioids that modulate nociceptive transmission. Exposure to a potentially infanticidal adult male rat suppresses pain-related behaviors in pre-weaning but not in older rats. This male-induced analgesia is mediated by l opioid receptors in the periaqueductal gray, a midbrain structure that is innervated by amygdala projections. To determine whether enkephalin, a l and d opioid receptor agonist, is activated by male exposure, mRNA levels of its precursor, preproenkephalin, were measured in subdivisions of the amygdala and the periaqueductal gray. In 14-day-old but not in 21-day-old rats, 5 min of male exposure induced analgesia to heat and increased preproenkephalin mRNA levels in the central nucleus of the amygdala but not in the periaqueductal gray. The change in the activation of enkephalinergic neurons in the central amygdala may contribute to the change in stress-induced analgesia during early ontogeny.

The present study examined the ability of LY235959, a competitive N-methyl-D-aspartate (NMDA) receptor antagonist, to attenuate behaviors and c-fos mRNA expression associated with acute morphine withdrawal in the infant rat. Rat pups were given a single dose of morphine (10.0 mg/kg, s.c.) or saline. Two hours later, pups were removed from the dam and injected with either LY235959 (10.0 mg/kg, s.c.) or saline. Fifteen minutes later acute morphine withdrawal was precipitated with naltrexone (10.0 mg/kg, s.c.) and behaviors were recorded every 15 s for the next 60 min. Immediately after behavioral testing, brain and spinal cord were assayed for c-fos mRNA analysis by solution hybridization. The intensity of the morphine withdrawal syndrome was reduced in pups pre-treated with LY235959. Withdrawal behaviors such as head moves, moving paws, rolling, and walking were decreased, and vocalizations were completely eliminated in pups pre-treated with LY2359559. Acute morphine withdrawal increased c-fos mRNA expression in the brain and the spinal cord, which was attenuated by pre-treatment of LY235959. Thus, in the 7-day-old rat, as in the adult, NMDA receptors play a role in the behavioral and molecular manifestations of acute morphine withdrawal.

This study investigated the effects of the NMDA receptor antagonist MK-801 on the development of morphine dependence in 7-, 14-, and 21-day-old rat pups. For 6.5 days, starting at 1, 8, or 15 days of age, rats were pretreated with MK-801 (0.03 or 0.1 mg/kg, bid) or saline; 15 min later, morphine sulfate (10 mg/kg) or saline was injected to induce opiate dependence. On the afternoon of the seventh day, pups were injected with MK-801 (0.1 mg/kg) or saline and 15 min later with naltrexone (1 mg/kg) to precipitate withdrawal. Pups were then placed in a warm chamber with the litter and their behavior scan-sampled every 15 sec for a total of 15 min. MK-801 failed to inhibit morphine withdrawal in the 7-day-old rat, but did attenuate the development of mor- phine dependence in both the 14- and 21-day-old rats. These results suggest that the NMDA receptor is not functionally active in opiate withdrawal until around the second to third week of postnatal life in the rat and that there exists a transition period for the NMDA receptor to play a role in the development of opiate dependence and withdrawal. Synapse 40:282-293, 2001. © 2001 Wiley-Liss, Inc.

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 (10.0 mg/kg) twice daily for 6.5 days. On the 7th day, pups were injected with NOS inhibitors (l-NAME or 7-NI), NMDA receptor antagonists (MK-801 or AP-5), or vehicle. After 15 min, the pups were injected with naltrexone (1 mg/kg) to precipitate withdrawal. Behavior for each pup was identified and recorded every 15 s for 10 min before naltrexone injection and 15 min 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.

Large numbers of human infants are exposed in utero to illicit opiate drugs such as heroin and to prescribed opiates such as methadone. It is therefore important to understand the effects of acute and chronic exposure to drugs such as these on the immature organism. A necessary step in understanding the long-term effects of early drug exposure is to describe fully how the drug acts on neurobehavioral systems in the neonate. Because infants typically experience withdrawal from opiates, either in utero or shortly after parturition, a fuller understanding of the consequences of opiate withdrawal in the immature animal is an important but largely unexplored area of research. Although there has been some question as to whether infants undergo withdrawal, more recent experiments demonstrate that the rat neonate experiences clear behavioral changes during opiate abstinence, including the induction of "dysphoric" states on cessation of drug exposure. It is proposed that the infant organism undergoes withdrawal but that the signs and symptoms experienced differ from those suffered by the adult.

This study investigated the effect of paraventricular nucleus (PVN) injection of neuropeptide Y (NPY) on milk and water intake of 2- and 15-day-old sated rats. On the day prior to testing, rat pups were removed from their mothers and implanted with a cannula directed unilaterally at the PVN. On the following day, each pup was implanted with an intra-oral cannula for oral infusion of milk or water that could be swallowed or rejected. Following a 1-hour period of satiation, each pup received a PVN injection of saline or a single dose of NPY (23.5–235.0 pmol). Milk or water intake was then assessed in a 1-hour test period. Injection of NPY into the PVN enhanced milk and water intake equally at 2 days of age. At 15 days, NPY produced a significantly greater enhancement of milk than water intake. These findings, which are similar to those observed previously with PVN injections of norepinephrine (NE), suggest that: (1) NPY receptors in the PVN, like α 2 -noradrenergic receptors, are functional very early in the postnatal development of the rat; (2) NPY, in addition to its orexigenic effect, produces a small but significant dipsogenic effect; and (3) NPY may function cooperatively with NE in the PVN to stimulate feeding and drinking beginning at a very early age.

The functional ontogeny of beta 2-adrenergic and dopaminergic receptors in the perifornical lateral hypothalamus (PLH) that mediate adrenergic and dopaminergic suppression of feeding in rats was investigated. Rat pups, ranging in age from 2 to 15 days, were removed from their mothers and implanted with a brain cannula directed unilaterally at the PLH or a more rostral site lateral to the anterior nucleus of the hypothalamus. On the next day, following a 22-h period of food and water deprivation, each pup was implanted with an intra-oral cannula for oral infusion of milk that could be swallowed or rejected. Subsequently, each pup received an intracerebral injection of saline, or a single dose of epinephrine (EPI, 0.1-30.0 nmol), the beta 2-adrenergic receptor agonist salbutamol (1.0-30.0 nmol) or the dopaminergic receptor agonist apomorphine (1.0-30.0 nmol). Milk intake was then assessed following a 1-h period of infusion. The results showed significant dose-dependent suppression of milk intake in pups as young as 2 days of age in response to PLH injection of EPI, salbutamol and apomorphine. In contrast to its effectiveness in the PLH at 2 days of age, EPI failed to suppress milk intake at this age following injection into a more rostral site lateral to the anterior nucleus of the hypothalamus. Together, these findings suggest that both beta 2-adrenergic and dopaminergic receptors, mediating adrenergic and dopaminergic suppression of feeding, are functionally mature very early in the postnatal development of the rat. Moreover, consistent with evidence in adult rats, these catecholaminergic receptors in young pups appear to be located in the region of the PLH.

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 agress with the reported conditioning of amphetamine's behavioral effects but differs in terms of the action on brain dopamine turnover.

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.

Male and female albino rats were tested for intraspecies aggression without the use of shock. In the first experiment, male pairs showed more biting attacks, offensive sideways movements, and self-grooming than did female pairs; male pairs also showed more stereotyped defensive/submissive behaviors and were wounded more frequently. The second experiment examined the effects of neonatal castration and testosterone propionate (TP) administration on fighting. Males castrated at birth attacked other males less frequently than did controls when tested with TP treatment as adults. The TP given at birth to neonatally castrated males restored attacks to control levels. Females given TP as neonates did not differ from either male or female controls. Other aggressive/defensive behaviors, however, did not show this pattern. The results suggest that while the presence of testosterone during a brief postnatal period and during adulthood is necessary for attack behavior to occur, other related behaviors may not be affected in a similar manner.

Previous studies have reported age-related changes in opiate receptors and in their response to narcotics during the process of normal growth of the brain. By inducing alterations in this developmental sequence, the present study attempted to provide correlates of the opiate receptor system with naloxone-induced anorexia. Offspring of mothers treated with morphine (7.5 mg/kg twice daily, s.c.) or saline during pregnancy, and infants from untreated mothers given morphine (5 mg/kg), naltrexone (10 mg/kg) or saline subcutaneously on postnatal days 1–5, were tested at days 10, 12 and 14 for deprivation-induced milk consumption following an acute dose of naloxone (1 or 5 mg/kg, i.p.). Naloxone reduced the food intake of 10- and 12-day old infants chronically treated with morphine postnatally. At age 14, naloxone reduced the food consumed by all the pretreatment groups, and pretreatment with morphine altered the dose-response curves for feeding modulation induced by naloxone. Naloxone had no effect on the food consumed by 10- or 12-day old offspring of mothers treated with morphine or saline, or on those age-groups that recieved naltrexone chronically or saline pretreatments postnatally. The observed changes occurred in the absence of gross malformations, drug-withdrawal symptoms and differences in activity. These results demonstrate that opiate receptors may participate in feeding.

To assess the effects of catecholaminergic drugs on independent feeding during development, preweanling rats were administered amphetamine (AMPH) or chlorpromazine (CPZ) and were allowed to ingest milk through anteriorly located intra-oral cannulas. In 1-hr milk-deprived rat pups, AMPH stimulated milk intake at 3, 7 and 10 days of age and suppressed intake at 15 days. In 22-hr-deprived pups, AMPH had no effect at 3, 7 and 10 days, but reliably suppressed intake at 15 days. CPZ stimulated intake in 3-, 10- and 15-day-old milk-satiated pups. In 22-hr-deprived pups, CPZ had no effect at 3 and 10 days, but stimulated intake at 15 days. While AMPH produced pronounced nonappetitive behavioral activation in conjunction with enhanced intake in 1-hr-deprived pups, AMPH-induced activation occurred without enhanced intake in 22-hr-derived pups. Thus, there was a dissociation between AMPH's effect on milk intake and AMPH-induced nonappetitive behavioral activation. Collectively, the present results support the following conclusions. First, a catecholaminergic system(s) that enhances independent feeding is present very early in postnatal development of the rat. Second, level of food deprivation is an important state-dependent variable when assessing the effects of AMPH and CPZ on independent feeding in preweanling rats.

The activity of dopamine- and norepinephrine-stimulated adenylate cyclase in hypothalamus and amygdala was studied during the estrous cycle of the female and following castration of the male rat. In the medial hypothalamus but not in the anterior hypothalamus or amygdala, stimulation of adenylate cyclase by dopamine was enhanced during proestrus and following castration. There were no changes in norepinephrine-stimulated adenylate cyclase in medial hypothalamus. Thus, an interaction may exist between gonadal hormones and dopamine receptors linked to adenylate cyclase in medial hypothalamus. The rapidity of the changes in receptor sensitivity suggests that this interaction plays an important role in physiological regulation.

Maintaining rats on a tryptophan-free diet for 4--6 days induced mouse killing in non-killer rats, and significantly facilitated killing in killer rats, as indicated by shorter latencies to kill the mice. The killing responses were similar in topography to the natural killing responses. These changes in killing behavior did not appear to be due to generalized changes in irritability. The increased killing after maintenance on a tryptophan-free diet was accompanied by a 26% reduction in brain serotonin (5-HT) and a 29% reduction in brain 5-hydroxyindoleacetic acid (5-HIAA). When the tryptophan-free diet was supplemented with L-tryptophan (0.5 or 2%), brain 5-HT and 5-HIAA were increased above control levels, and the rat's killing response appeared normal both in terms of latency and topography, similar to that seen in control chow fed animals. While rats consumed less of the tryptophan-free and tryptophan supplemented diets, control subjects deprived of chow such that they lost as much weight as rats fed the tryptophan-free diet, did not show changes in killing behavior. These results are consistent with the hypothesis that central serotonergic systems exert inhibitory control over mouse killing behavior in rats.

Adrenalectomy affects analgesic responses differentially, potentiating morphine and cold-water swim analgesia while reducing prolonged, intermittent foot shock analgesia. Adrenocortical influences also exhibit differential effects: dexamethasone pretreatment potentiates morphine analgesia while reducing both forms of stress-induced analgesia. In assessing sympathomedullary influences in analgesic processes, adrenal demedullation reduces prolonged, intermittent foot shock analgesia. The present study examined the effects of adrenal demedullation both with (MADX/6-OHDA) and without (MADX) peripheral 6-hydroxydopamine injections upon the analgesic responses following 2°C and 15°C swims and following 5 mg/kg and 2.5 mg/kg doses of morphine. Significant reductions were observed in adrenal epinephrine levels in all MADX rats and in heart norepinephrine levels in MADX/6-OHDA rats. Despite the adrenal demedullation and sympathetic dennervation, neither the MADX group nor the MADX/6-OHDA group differed significantly from sham control rats in their baseline post-operative or post-injection flinch-jump thresholds, in their analgesic responses following the 2°C and 15°C swims, or in their analgesic responses following the 5 mg/kg and 2.5 mg/kg morphine doses. These data are assessed in terms of divergent adrenal influences upon various analgesic responses.