Ilona Vathy

Hormonal Activation of Female Sexual Behavior is Accompanied by Hypothalamic Norepinephrine Release

The present study employed the intracranial microdialysis technique to measure norepinephrine release in the ventrolateral dendritic fields of the ventromedial hypothalamus of freely‐moving animals before and during ovarian steroid (estradiol and progesterone) activation of female sexual behavior (lordosis). One day after implantation of a dialysis probe, animals were injected with 3 μg of estradiol benzoate followed 44 h later by 200 μg of progesterone. Introduction of a male rat 4 h after progesterone treatment was correlated with dramatic increases in extracellular norepinephrine levels measured in dialysates of the ventrolateral ventromedial hypothalamus of female rats which displayed high levels of lordosis behavior. In contrast, female rats given the same steroid treatment but which did not show lordosis responses did not have elevated norepinephrine levels in their dialysates. Moreover, animals that received an estrogen antagonist concurrently with the estrogen treatment had neither an increase in ventromedial hypothalamic levels of norepinephrine during behavior testing nor did they display lordosis. These results indicate a close relationship among ovarian steroids, noradrenergic transmission in the ventromedial hypothalamus, and the expression of female sexual behavior.

Effects of prenatal morphine on adult sexual behavior and brain catecholamines in rats

Female rats exposed to morphine in utero (5-10 mg/kg twice a day on days 11-18 of gestation) were significantly inhibited in their sexual behavior when compared to saline-exposed controls. In contrast, males exposed prenatally to morphine had shorter post-ejaculatory intromission latencies and, after the first test, exhibited increased mounting and intromitting activity relative to controls. Examination of the brain catecholamine content revealed that morphine in utero may permanently alter adult hypothalamic norepinephrine levels in male and female rats. The morphine-induced alterations in hypothalamic norepinephrine levels were sexually dimorphic. In the hypothalamus of male rats, norepinephrine content was increased 95%, whereas in the hypothalamus of female rats it was decreased 57% relative to controls. These results suggest that prenatal morphine exposure, which differentially affects adult male and female sexual behavior, also alters hypothalamic norepinephrine content in a sexually dimorphic fashion.

Prenatal morphine exposure differentially alters learning and memory in male and female rats.

The present study tested the hypothesis that exposure to morphine on prenatal days 11-18 impairs performance on tasks requiring learning and memory in adult male and female rats. In Experiment 1, a symmetrical maze was used to measure learning. In Experiment 2, an eight-arm radial maze was used to assess working spatial memory. The results of Experiment 1 demonstrated that prenatal morphine exposure reduces the time needed to complete the trials, but does not affect the accuracy of performance in male rats. In contrast, prenatal drug treatment had no effects on either the time or the accuracy of performance in female rats. In Experiment 2, both male and female morphine-exposed rats needed more time to complete regular trials (no delay) than controls; however, morphine exposure in male rats did not affect performance on tasks requiring memory, measured with delay trials, but hindered it in ovariohysterectomized (OVX) female rats. In OVX females, replacement injections of both estrogen and progesterone restored the impairment of performance on delay trials produced by prenatal morphine exposure. Thus, the present study demonstrates that prenatal morphine exposure differentially alters performance of adult male and female rats on tasks requiring learning and spatial memory.

Prenatal opiate exposure: long-term CNS consequences in the stress system of the offspring

Our data show that prenatal morphine exposure induces long-term alterations in adult brain and behavior in both male and female rats, and these alterations are sex-specific. It is also evident that the alterations are not restricted to a single brain site or to a single neurotransmitter or neuromodulator. Moreover, there are long-term alterations in both the norepinephrine (NE) and opioid systems in several brain regions involved in stress responses and in the maintenance of homeostatic balance between the external environment, the brain and the rest of the body. Thus, this short paper reviews the prenatal morphine exposure data and highlights gaps in stress response to drug vulnerability/predisposition as an adult.

Repeated morphine administration during pregnancy attenuates maternal behavior

The present study tested the hypothesis that repeated administration of morphine on days 11-18 of pregnancy alters maternal behavior. Saline- and morphine-treated mothers were observed with their pups in two experiments. Rats were always tested twice a day during the light and dark phases of the reverse light/dark cycle. In Experiment 1, 12 types of activities and three types of nursing positions of mothers were recorded ten times during each 50-minute session for the 23-day lactation period. A decrease in nursing and active maternal behavior, and an increase in self-care, rearing and sniffing was found in morphine-treated mothers. Additionally, both saline- and morphine-treated mothers exhibited significantly more maternal behavior during the light, and non-maternal activities during the dark sessions of each day. Moreover, both saline- and morphine-treated mothers displayed significantly less maternal behavior and more non-maternal activities as postpartum time progressed. In Experiment 2, a different group of mothers was tested for pup retrieval from postnatal days 1 through 12. Morphine-treated mothers were slower than saline-treated mothers in retrieving all pups into the nest. However, there were no differences in latency to carry the first pup and return him/her to the nest. No unusual maternal behaviors were observed during the retrieval tests. Thus, the present study suggests that morphine administration during the second half of pregnancy attenuates some components of maternal behavior and increases non-maternal activities of mothers.

Prenatal exposure to morphine alters brain μ opioid receptor characteristics in rats

Prenatal morphine exposure alters neither the binding capacity nor the affinity of ligand binding to mu opioid receptors of adult male brains. However, males have significantly higher Bmax in the hypothalamus than ovariectomized females. In females, prenatal exposure to morphine reduces the Bmax of mu opioid receptors 25% in the hypothalamus and preoptic area. Estrogen treatment increases the Bmax of mu opioid receptors in the striatum of all ovariectomized females but in the hypothalamus only of morphine-exposed females, thereby eliminating the sex difference observed in control animals.

Modulation of catecholamine turnover rate in brain regions of rats exposed prenatally to morphine

The concentration and turnover rate of brain catecholamines were measured in the hypothalamus, preoptic area (POA), frontal cortex, striatum and cerebellum of adult male and female rats exposed in utero to morphine (5-10 mg/kg/twice a day) during gestation days 11-18. Norepinephrine (NE) and dopamine (DA) turnover rates were estimated following alpha-methylparatyrosine (AMPT) administration. Prenatal morphine altered NE content and turnover in male and female rats in a regionally specific, sexually dimorphic manner. Basal NE content increased approximately 60% in the hypothalamus of male rats, but it decreased about 30% in the hypothalamus of female rats. NE turnover in the hypothalamus of morphine-exposed rats increased 50% in males and decreased 50% in females. Prenatal morphine had no effects on NE turnover in the male POA, but in female rats NE turnover decreased approximately 60%. Alterations in the frontal cortex of morphine-exposed male and female rats resembled the pattern in the hypothalamus; however, the differences did not reach statistical significance. In addition, prenatal morphine had no effect on striatal or cerebellar NE or on basal levels or turnover of DA in any brain regions. These results demonstrate that prenatal morphine alters the content and turnover of NE in a sexually dimorphic manner in specific brain regions of male and female rats, suggesting alterations in the density of terminals and/or utilization of NE. These sexually dimorphic alterations in hypothalamic NE induced by prenatal morphine may be related to the changes observed in adult male and female sexual behavior in our previous work.

Sexually dimorphic effects of prenatal cocaine on adult sexual behavior and brain catecholamines in rats.

Exposure to cocaine in utero (10 mg/kg twice a day on days 11-18 of gestation) differentially altered adult sexual behavior, brain catecholamine levels and perhaps open field activity in male and female rats. Female rats exposed to cocaine were significantly inhibited in their sexual behavior when compared to saline-exposed controls. In contrast, males exposed prenatally to cocaine exhibited normal or even facilitated sexual behavior. In particular, the significantly shorter post-ejaculatory intromission intervals displayed by cocaine-exposed males indicated that these animals might have enhanced sexual arousal or motivation. In tests for open field activity, males consistently showed higher levels of rearing behavior than females. Prenatal cocaine tended to reduce rearing in females, but this effect was not significant. Cocaine-exposed male rats had significantly higher norepinephrine and dopamine levels in the preoptic area than saline-exposed controls, whereas cocaine did not affect catecholamine content in male hypothalamus, striatum, cortex or cerebellum. Cocaine- and saline-exposed females had similar catecholamine levels in all brain regions. These results suggest that exposure of developing animals to modest doses of cocaine during mid to late gestation results in long-lasting, sexually dimorphic alterations of adult sexual behavior and brain catecholamines in rats.

Quantitative peptidomics in mice: Effect of cocaine treatment

We recently developed a quantitative peptidomics method using stable isotopic labels and mass spectrometry to both quantify and identify a large number of peptides. To test this approach and screen for peptides regulated by cocaine administration, 32 Cpefat/fat mice and 16 wild-type mice were treated twice daily for 5 d either with saline or 10 mg/kg cocaine. Peptides were extracted from striatum, hypothalamus, hippocampus, and trimethylammonium butyrate containing either nine deuterium or nine hydrogen atoms. Pools of heavy- and light-labeled peptides were combined, purified on an anhydrotrypsin affinity column, and analyzed on a reverse-phase column coupled to an electrospray ionization quadrapole time-of-flight mass spectrometer. Changes in peptide levels upon cocaine treatment were determined from the relative peak intensities of the cocaine versus saline peaks, and peptides were identified from collision-induced dissociation spectra. Ten peptides were found to increase or decrease in each of two separate analyses from distinct groups of mice. Peptides found to increase corresponded to fragments of proenkephalin, prothyrotropin-releasing hormone, provasopressin, proSAAS, secretogranin II, chromogranin B, and peptidyl-glycine-α-amidating mono-oxygenase in the hypothalamus. The same peptidyl-glycine-α-amidating mono-oxygenase peptide decreased in the prefrontal cortex, along with striatal neurokinin B and two unidentified peptides. Thirty other peptides were not substantially affected by cocaine treatment in both replicates. Taken together, the quantitative peptidomics approach provides an efficient method to screen for changes in a large number of peptides.

Prenatal exposure to morphine differentially alters gonadal hormone regulation of δ-opioid receptor binding in male and female rats

The present study tested the hypothesis that exposure to morphine on gestation days 11-18 differentially alters delta-opioid receptors in the brain of adult male and female rats. In Experiment 1, the binding characteristics of delta-opioid receptors were examined in membrane homogenates from six brain regions, including the hypothalamus (HYP), preoptic area, frontal cortex (CX), ventral tegmental area, striatum (STR) and cerebellum of adult male and female rats. In Experiment 2, the density of delta-opioid receptors was assessed in the CX and STR using receptor autoradiography. Prenatal morphine exposure has no effects on delta-opioid receptors in the brain of gonadally intact, adult male rats regardless of methodology. However, when male rats were gonadectomized in Experiment 2, morphine-exposed males have fewer delta-opioid receptors than controls in the CX but not in the STR. These reductions in cortical delta-opioid receptors are restored by testosterone replacement, demonstrating that prenatal morphine exposure alters testosterone regulation in the CX of male rats. In ovariectomized (OVX) female rats, prenatal morphine exposure increases the density of delta-opioid receptors in the frontal CX. Interestingly, this up-regulation of delta-opioid receptors is not present when the CX is investigated by autoradiography. Moreover, progesterone given alone or in combination with estrogen reduces the density of delta-opioid receptors in the CX and STR of both saline- and morphine-exposed, OVX females. Thus, mid to late gestational morphine exposure differentially alters the influence of adult gonadal hormones on delta-opioid receptors in the CX, decreasing the sensitivity in females and increasing it in males. This is also the first report to demonstrate that gonadal hormones regulate delta receptor densities in brain regions other than the HYP of OVX females.