Lori L. Badura

Male-induced estrus synchronization in the female Siberian hamster (Phodopus sungorus sungorus)

Olfactory cues play an integral role in the organization of events that mediate reproductive success. In a variety of species, priming pheromones, in particular, are important for ensuring reproductive fitness. To date, very little research has focused on how male-emitted priming pheromones, such as those that regulate the onset of puberty and estrus synchronization in females, affect the reproductive physiology of the female Siberian hamster (Phodopus sungorus sungorus). This lack of research may be due to the physiology of the Phodopus genus; vaginal cytology cannot be used as a reliable indicator of estrus or ovulation. Using a jugular cannulation technique to determine estrous stage by blood analysis of prolactin and luteinizing hormone, we sought to determine if male priming pheromones affect estrous cyclicity in the female Siberian hamster and, if so, whether the production of these priming pheromones is androgen dependent. Our results showed that females exposed to bedding from mature, intact males showed a significantly higher incidence of proestrus 3 days later than did females exposed to the bedding of mature, gonadectomized males. Therefore, we found that not only do male Siberian hamsters emit chemical signals that induce estrus synchronization, but also that this ability is likely to be androgen dependent.

Chronic administration of sodium valproic acid slows pubertal maturation in inbred DBA/2J mice: Skeletal, histological, and endocrinological evidence

Sodium valproic acid (VPA) has been reported to occasionally delay pubertal maturation in children. In the current study, we sought to establish a valid animal model with which to further investigate the neuroendocrinological sequelae of VPA administration. Male and female DBA/2J mice were weaned at 2 weeks of age and administered either VPA (17-20 mg/kg/day) or control solution via drinking water. Animals were weighed and sacrificed via decapitation at 4, 6, or 8 weeks of age. Testes and ovaries were prepared for histological analyses. In addition, the length of the left humerus bone from each animal was obtained as an index of skeletal growth, and trunk blood was assayed for circulating follicle-stimulating hormone (FSH) and prolactin (PRL). For males, testicular weights of the animals receiving VPA were significantly lower than those of control animals at all three sampling ages. No between-group differences were found for body weight at any sampling age, and yet the rate of skeletal maturation (as indexed by humerus length) was decreased significantly for the VPA-treated males at all three sampling periods. Additionally, while hormone levels did not consistently differ, histological analyses of the gonadal tissue demonstrated significantly decreased rats of spermatogenesis at all sampling points for VPA-treated animals. For females, chronic VPA administration led to a significant reduction in uterine weight at the 4 and 6 week sampling periods, and yet by 8 weeks of age the uterine weights for the two groups did not differ.(ABSTRACT TRUNCATED AT 250 WORDS)

GABA levels within the medial preoptic area: effects of chronic administration of sodium valproic acid

Sodium valproic acid (VPA) is a widely prescribed anticonvulsant medication that has been shown to interfere with pubertal maturation of the reproductive system, and induce endocrine abnormalities in adults, within a subset of the clinical population. While VPAs mechanism of action is still poorly understood, it may exert its anti-reproductive effects by enhancing GABAergic inhibition of the GnRH neuronal population within the medial preoptic area (mPOA). The purpose of this study was to determine if chronic administration of VPA alters GABA levels within the mPOA region. In Experiment 1, the mPOA, caudate, and arcuate nucleus regions were harvested from VPA-treated and control mice. Analysis of whole tissue content of GABA revealed that levels were lower in the caudate and arcuate nucleus regions of VPA-treated animals, whereas there were no group differences for the mPOA region. Collapsing across drug group, there was also a trend for males having overall higher levels of GABA as compared to females. In Experiments 2 and 3, mice were implanted with microdialysis probes within the mPOA region and sampled for extracellular GABA levels. Females (Exp. 3) were sampled either on diestrous, proestrous, or estrous. Results from males (Exp. 2) revealed that VPA enhanced extracellular GABA levels in the mPOA region compared with controls. However, GABA levels for both groups remained stable across the sampling period. Conversely, in Exp. 3, females showed cyclical release of GABA across the sampling period. For control females, GABA levels increased during the afternoon on all cycle days, but the rise on proestrus was smaller than on other cycle days. VPA-treated animals showed an overall reduction in GABA levels compared with controls. Furthermore, while GABA increased over sampling time on estrus and diestrus days of the cycle, there was not a significant rise in GABA on proestrus. These data indicate: (1) regional specificity in VPA effects upon GABA levels, (2) a sex difference in the effects of VPA on GABA levels within the mPOA, and (3) GABA levels increase on the afternoon of all days of the estrous cycle with VPA attenuating the rise seen on the afternoon of proestrus. These results provide evidence that VPA effects upon the reproductive axis may involve changes in GABA release, and that males and females show different patterns of neurochemical response to the drug.

A potential mechanism of slowed pubertal maturation after chronic administration of sodium valproic acid

Sodium valproic acid (VPA) occasionally delays pubertal maturation in children and gonadal and skeletal growth in juvenile, seizure-prone, inbred DBA/2J mice. Fourteen-day-old mice received either VPA or control solution, and the medial preoptic area (mPOA) of the hypothalamus was studied after 4, 7, 10, 14, or 18 days by immunocytochemically processing for gonadotropin-releasing hormone (GnRH). Significant differences were found for the proportion of bipolar to unipolar neurons in the mPOA. VPA may slow pubertal maturation by altering neurochemical systems that normally play an important role in timing maturation of the GnRH pulse generator.

5HT and 5HIAA dialysate levels within the arcuate nucleus of the hypothalamus: relationship with photoperiod-driven differences in serum prolactin and luteinizing hormone in the Siberian hamster

This study examined the relationship between dialysate levels of serotonin (5HT), and its major metabolite 5HIIAA within the arcuate nucleus of the hypothalamus (ARC) and serum gonadotropin levels under two different in vivo paradigms. Experiment 1 evaluated the relationship between dialysate levels of 5HT and 5HIAA within the ARC and circulating prolactin (PRL) and lutenizing hormone (LH) levels under long- and short-day photoperiod conditions. In experiment 2, the profile of 5HT and 5HIAA dialysate levels within the ARC on the afternoon of proestrous was investigated to determine if changes in serotonergic neurotransmission are correlated with preovulatory surges in LH and PRL. Adult male and female Siberian hamsters were housed either in long-day (16L:8D) or short-day (10L:14D) photoperiods for 8 weeks. Dialysis samples were collected every hour for 5 h (12.00-17.00 h) and blood samples were collected via a jugular cannula every hour for analysis of LH and PRL levels. ARC 5HT and 5HIAA dialysate levels were significantly higher in short-day exposed female hamsters, correlating with suppressed basal LH and PRL secretion when compared to their long-day counterparts. Short-day housed male hamsters displayed significantly higher dialysate levels of 5HIAA than males exposed to a long-day photoperiod-5HT was below the lower limit of detection regardless of photoperiod exposure. Long-day females in proestrus showed no change in dialysate levels of 5HT or 5HIAA within the ARC just prior to the onset of the afternoon surge of LH and PRL. Our results indicate that elevated 5HT and 5HIAA dialysate levels within the ARC may regulate photoperiod effects upon LH and PRL secretion, but not the preovulatory surges of LH and PRL.

Sodium valproate alters GnRH-GABA interactions during development in seizure-prone mice.

During reproductive maturation, characteristic changes occur in the morphology of the gonadotropin releasing hormone (GnRH) cell population within the hypothalamus. In the early stages of development, GnRH neurons are bipolar cells; however, just before pubertal onset, the majority of these neurons transform into unipolar cells. Our laboratory has reported that valproic acid (VPA), an antiepileptic medication that has previously been shown to slow the velocity of pubertal development in both humans and seizure-prone mice, is capable of delaying the normal process of GnRH morphological differentiation. As VPA is primarily believed to act via a GABAergic mechanism, the present study investigated potential influences of VPA on GnRH-GABA interactions within the medial preoptic area (mPOA) across pubertal development (experiment 1), as well as in adult animals (experiment 2). The results from experiment 1 revealed the expected drug effects on GnRH cell morphology. For VPA animals, there was a greater percentage of bipolar neurons at every time period except for the 24-day sample. Additionally, VPA animals had greater numbers of bipolar and unipolar GnRH neurons with GABA associations across all ages. However, experiment 2 showed a lack of drug effects on GnRH-GABA interactions in adulthood. These results suggest that VPA may delay GnRH cell morphological maturation by altering the density of GABAergic inputs to GnRH neurons. These inputs may normally play a role in timing the activation of the GnRH pulse generator. However, any neuroendocrine effects of VPA in adulthood are most likely due to the actions of VPA at another level of the hypothalamic-pituitary-gonadal axis.

Noradrenergic regulation of prolactin secretion at the level of the paraventricular nucleus of the hypothalamus: functional significance of the alpha-1b and beta-adrenergic receptor subtypes.

Previous research has demonstrated that in the Siberian hamster, both photoperiod and estrous cyclicity alter the profile of noradrenergic activity with the paraventricular nucleus of the hypothalamus (PVN), and that noradrenergic activity is correlated with changes in circulating levels of prolactin. Work from our laboratory has demonstrated an inhibitory role for norepinephrine (NE) acting at the alpha-2 receptor subtype within the PVN on serum prolactin levels; however, the functional significance of other adrenergic receptor subtypes on this system is unknown. The purpose of this study was to investigate the functional significance of the alpha-1b and beta-adrenergic receptor subtypes at the level of the PVN on circulating levels of prolactin. These experiments were performed in male Siberian hamsters using reverse microdialysis coupled with serial blood sampling. In Experiment 1, infusion of l-phenylephrine hydrochloride (alpha-1b agonist) initiated a dose-dependent increase in circulating prolactin, whereas infusion of chloroethylclonidine (alpha-1b antagonist) induced a significant dose-dependent decline in prolactin. In Experiment 2, intraparaventricular administration of propranolol (beta antagonist) initiated a significant increase in prolactin levels in a dose-dependent manner, whereas isoproterenol (beta agonist) induced a dose-dependent decline in prolactin. The results of this study indicate that both the alpha-1b and beta-adrenergic receptor subtypes have a significant role in regulating circulating levels of prolactin at the level of the PVN in the Siberian hamster.

Infusion of Alpha-2-Adrenergic Agents into the Paraventricular and Arcuate Nuclei of the Hypothalamus in the Siberian Hamster: Opposing Effects on Basal Prolactin

Although the α2-adrenergic receptor subtype has been shown to have a significant influence on circulating levels of prolactin (PRL), its exact role remains unclear. A multitude of studies have demonstrated that blockade of the α2-receptor can either elevate or decrease circulating levels of PRL. α2-Receptor-mediated control of both stimulatory and inhibitory arms of the PRL regulatory system may explain this discrepancy. Activation of the α2-receptor has been shown to inhibit the activity of its target cell, and therefore antagonism of the α2-receptor within a stimulatory component (e.g., paraventricular nucleus (PVN) of the hypothalamus) would theoretically have the opposite effect that it would have within an inhibitory component (arcuate nucleus of the hypothalamus). Thus, the purpose of this study was to investigate the functional role of the α2-adrenergic receptor in modulating circulating levels of PRL both at the level of the PVN and arcuate using reverse microdialysis of α2-adrenergic agents coupled with serial blood sampling in the male Siberian hamster. Male hamsters were fitted with a jugular cannula for serial blood sampling, and an indwelling microdialysis probe for intrahypothalamic drug administration between 08:00 and 10:00 h. Blood samples were collected every hour for 5 h (12:00–17:00 h). During the third sampling period, atipamezole (α2-antagonist) or medetomidine (α2-agonist) at one of three doses were infused into the PVN or the arcuate to assess effects on basal PRL. At the level of the PVN, infusion of atipamezole initiated an increase in basal PRL in a dose-dependent fashion, whereas infusion of medetomidine induced a significant decline in basal PRL in a dose-dependent fashion. In the arcuate, only the highest dose of atipamezole had an effect on PRL, and this was in the opposite direction from that seen in the PVN. Infusion of medetomidine did not have a significant effect on basal PRL levels; however, a trend toward a significant elevation was observed for the highest dose. These results suggest that the α2-receptor subtype may have opposite effects on circulating levels of PRL within the PVN and arcuate regions, and may explain why antagonism of the α2-receptor has been shown to initiate both surges and declines in basal levels of PRL.

Differential prolactin responsiveness to stress in left- and right-pawed mice

Behavioral laterality has been shown to be related to immunological and endocrinological responsiveness in mice. The current study extended these findings by examining the pituitary hormone prolactin, and its response to stress, in left- and right-pawed mice. In two experiments, mice were tested on two measures of behavioral laterality, the Collins paw preference test and the Lateral Paw Preference (LPP) test. Circulating PRL levels were then measured in all subjects under baseline and stressed conditions. In Experiment 1, the PRL stress response was related to Collins paw preference in interaction with coat-color among females of a genetically heterogeneous group. Left-pawed black females showed a significant increase in PRL, while right-pawed black females did not. Among agouti mice, left-pawed females showed a significant decrease, while right-pawed did not. Experiment 2 used a genetically homogeneous group of agouti C3H mice. Again there were effects of Collins paw preference among females, with the right-pawed showing a significant increase in PRL following stress. In neither experiment were there effects of LPP. In neither experiment were there lateralized differences among males. These results indicate lateralized regulation of PRL among females, similar to that previously observed for ACTH regulation.

Norepinephrine Dialysate Levels in the Hypothalamic Paraventricular Nucleus: Influence on Photoperiod-Driven Prolactin Levels in the Female Siberian Hamster

Basal prolactin (PRL) levels in Siberian hamsters are modulated by ambient photoperiod via the nocturnal melatonin signal. Recent evidence from our laboratory has demonstrated that norepinephrine (NE), a putative neurochemical regulator of PRL secretion shows photoperiod-dependent fluctuations in the hypothalamic paraventricular nucleus (PVN) that are independent of the pineal melatonin signal. NE content in the PVN is elevated under a short-day photoperiod with a time course that follows the short-day-induced decline in PRL. NE could thus modulate the release of a prolactin-releasing factor that subsequently drives PRL synthesis and release rates. In order to determine whether NE release in the PVN correlates with basal PRL levels, this study evaluated the relationship between extracellular NE levels in the PVN and basal PRL under long- and short-day photoperiod conditions. In addition, the profile of NE during the proestrous surge of PRL was investigated. Female Siberian hamsters were housed either in long-day or short-day photoperiods for 8 weeks. After 6 weeks of photoperiod exposure, the animals were implanted with a stainless steel guide cannula aimed at the PVN. Two weeks later, the animals were fitted with a jugular cannula for serial blood sampling and implanted with an indwelling microdialysis probe. Dialysis samples were collected every 20 min for 5 h (12.00–17.00 h) from short-day-exposed animals and from long-day-exposed animals classified as being either in diestrus or proestrus. Blood samples were collected every hour and analyzed for PRL levels by radioimmunoassay. NE and methoxy-4-hydrophenylglycol (MHPG) levels were significantly higher in short-day-exposed animals, correlating with the suppressed basal secretion of PRL. Both long-day groups had lower NE and MHPG levels, and higher PRL, than the short-day group. However, long-day animals showed a significant decline in NE in proestrus just prior to the onset of the afternoon surge of PRL. These data suggest that NE released within the PVN participates in the regulation of photoperiod effects upon PRL secretion, as well as that of the afternoon surge of PRL on proestrus.