James Janik

Orphanin FQ stimulates prolactin and growth hormone release in male and female rats

Intracerebroventricular administration of Orphanin FQ (5.5, 55 or 550 pmol) caused a dose-related increase in prolactin secretion in both male and female rats and stimulated GH secretion in males. The magnitude of the prolactin secretory response was greater in females than in males. These effects of OFQ on prolactin and growth hormone release are the same as the stimulatory effects of the endogenous opioid peptides.

Immunoneutralization of Endogenous Opioid Peptides Prevents the Suckling-Induced Prolactin Increase and the Inhibition of Tuberoinfundibular Dopaminergic Neurons

Previous studies have shown that the endogenous opioid peptides, acting at specific opiate receptor subtypes, are involved in the suckling-induced prolactin secretory response. The prolactin increase elicited by suckling is due, at least in part, to an inhibition of tuberoinfundibular dopaminergic (TIDA) neurons in the hypothalamus. We investigated the effects of immunoneutralization of dynorphin, leu-enkephalin and met-enkephalin on the suckling-induced prolactin increase and on the activity of the TIDA neurons in lactating female rats between days 7 and 12 postpartum. Rats were injected into the right lateral ventricle with antiserum specific for one of these three peptides. Control rats were administered equal amounts of immunoglobulin proteins. Suckling produced a profound and significant increase in prolactin levels, as well as a decrease in DOPA accumulation in the median eminence of lactating rats. Administration of immunoglobulin concentrations of up to 3.6 µg did not inhibit the prolactin secretory response to the suckling stimulus and did not prevent the suckling-induced inhibition of TIDA neurons. Antisera to all three endogenous opioid peptides abolished the suckling-induced prolactin increase and prevented the inhibition in DOPA accumulation in the median eminence. Thus, the endogenous opioid peptides, dynorphin, leu-enkephalin and met-enkephalin, are essential for the prolactin secretory response to suckling and inhibition of TIDA neuronal activity is at least one of the mechanisms of action utilized by these peptides.

Estrogen regulation of neurotrophin expression in sympathetic neurons and vascular targets

We hypothesize that estrogen exerts a modulatory effect on sympathetic neurons to reduce neural cardiovascular tone and that these effects are modulated by nerve growth factor (NGF), a neurotrophin that regulates sympathetic neuron survival and maintenance. We examined the effects of estrogen on NGF and tyrosine hydroxylase (TH) protein content in specific vascular targets. Ovariectomized, adult Sprague-Dawley rats were implanted with placebo or 17beta-estradiol (release rate, 0.05 mg/day). Fourteen days later, NGF levels in the superior cervical ganglia (SCG) and its targets, the heart, external carotid artery, and the extracerebral blood vessels, as well as estrogen receptor alpha (ERalpha) content levels in the heart, were determined using semi-quantitative Western blot analysis. TH levels in the SCG and extracerebral blood vessels were determined by Western blotting and immunocytochemistry, respectively. Circulating levels of 17beta-estradiol and prolactin (PRL) were quantified by RIA. Estrogen replacement significantly decreased NGF protein in the SCG and its targets, the external carotid artery, heart and extracerebral blood vessels. TH protein associated with the extracerebral blood vessels was also significantly decreased, but ERalpha levels were significantly increased in the heart following estrogen replacement. These results indicate that estrogen reduces NGF protein content in sympathetic vascular targets, which may lead to decreased sympathetic innervations to these targets, and therefore reduced sympathetic regulation. In addition, the estrogen-induced increase in ERalpha levels in the heart, a target tissue of the SCG, suggests that estrogen may sensitize the heart to further estrogen modulation, and possibly increase vasodilation of the coronary vasculature.

Effects of energy status and diet on Bdnf expression in the ventromedial hypothalamus of male and female rats.

Sex differences exist in the regulation of energy homeostasis in response to calorie scarcity or excess. Brain-derived neurotrophic factor (BDNF) is one of the anorexigenic neuropeptides regulating energy homeostasis. Expression of Bdnf mRNA in the ventromedial nucleus of the hypothalamus (VMH) is closely associated with energy and reproductive status. We hypothesized that Bdnf expression in the VMH was differentially regulated by altered energy balance in male and female rats. Using dietary intervention, including fasting-induced negative energy status and high-fat diet (HFD) feeding-induced positive energy status, along with low-fat diet (LFD) feeding and HFD pair-feeding (HFD-PF), effects of diets and changes in energy status on VMH Bdnf expression were compared between male and female rats. Fasted males but not females had lower VMH Bdnf expression than their fed counterparts following 24-hour fasting, suggesting that fasted males reduced Bdnf expression to drive hyperphagia and body weight gain. Male HFD obese and HFD-PF non-obese rats had similarly reduced expression of Bdnf compared with LFD males, indicating that dampened Bdnf expression was associated with feeding a diet high in fat instead of increased adiposity. Decreased BDNF signaling during HFD feeding would increase a drive to eat and may contribute to diet-induced obesity in males. In contrast, VMH Bdnf expression was stably maintained in females when energy homeostasis was disturbed. These results suggest sex-distinct regulation of central Bdnf expression by diet and energy status.

Opiate Receptor Subtype Involvement in the Stimulation of Prolactin Release by β-Endorphin in Female Rats

The prolactin secretory response to beta-endorphin and the involvement of opiate receptor subtypes in this response was determined in both diestrous and postpartum, lactating female rats. The involvement of the mu-, delta- and/or kappa-site was determined by administering specific antagonists for each of these sites prior to beta-endorphin. beta-Funaltrexamine (beta-FNA, 1 or 5 micrograms) was administered to block mu-sites, ICI 154,129 (5, 10 or 25 micrograms) blocked delta-sites and nor-binaltorphimine (norBNI, 8 micrograms) blocked kappa-sites. The ability of beta-FNA and ICI 154,129 to block prolactin secretion following morphine administration was also determined. A dose response study for beta-endorphin indicated that beta-endorphin, at doses as low as 25 ng, was a potent stimulus for prolactin release producing an increase in prolactin that mimicked the suckling-induced prolactin increase. In addition, all three antagonists were capable of antagonizing the stimulatory effect of beta-endorphin in both diestrous and postpartum female rats. These results indicate that beta-endorphin is a potent stimulus for prolactin secretion and that these three opiate receptor subtypes interact to produce its stimulatory effect on prolactin release.

Effects of immobilization stress on tuberoinfundibular dopaminergic (tida) neuronal activity and prolactin levels in lactating and non-lactating female rats

The effects of immobilization stress on the prolactin secretory response and on the activity of the tuberoinfundibular dopaminergic (TIDA) neurons were determined in intact, virgin female rats on the morning of diestrus or proestrus and in post-partum, lactating female rats. The virgin females exhibited a significant increase in circulating levels of prolactin which was evident by 1 minute and persisted during the immobilization (5 minutes). In contrast, the prolactin secretory response in lactating females was significantly attenuated compared to non-lactating animals. The activity of the TIDA neurons was not altered by the 5 minutes of stress. Even after 30 minutes of immobilization, TIDA neuronal activity was not affected in either the lactating or cycling females. These data suggest that the cycling female rat is capable of a prolactin secretory response to the stressor without inhibition of TIDA neuronal activity. It seems likely that prolactin releasing factors mediate this response. In contrast, stress did not produce a similar prolactin increase during lactation. It seems likely that, during lactation, the pituitary is not sensitive to releasing factors unless the TIDA neurons are inhibited. There appear to be differences in the sensitivity of the pituitary depending on the physiological state of the model employed.

IMMUNONEUTRALIZATION OF β-ENDORPHIN BLOCKS PROLACTIN RELEASE DURING SUCKLING WITHOUT AFFECTING TUBEROINFUNDIBULAR DOPAMINERGIC NEURAL ACTIVITY

The effect of immunoneutralization of beta-endorphin on the suckling-induced prolactin increase and on the activity of the tuberoinfundibular dopaminergic (TIDA) neurons was determined in lactating female rats between days 8 - 12 post-partum. Two antisera were used in the immunoneutralization studies. Both were specific for beta-endorphin, exhibiting little cross reactivity with met- or leu-enkephalin or dynorphin. Antisera to beta-endorphin completely abolished the suckling-induced prolactin increase indicating that this endogenous opioid peptide is involved in this response. Suckling significantly inhibited DOPA accumulation in the median eminence and antiserum to beta-endorphin did not prevent this inhibition. Additionally, 5-endorphin antiserum significantly reduced TIDA neural activity even in pup-deprived dams. These results indicate that beta-endorphin is involved in the prolactin secretory response to suckling but that inhibition of TIDA neuronal activity is not its mechanism of action. Other possible mechanisms are discussed.

Morphine induced analgesia is attenuated in post-partum lactating rats

The analgesic effects of morphine administration were determined in post-partum, lactating female rats, as well as in intact, cycling females during the diestrous stage of the estrous cycle. All doses of morphine (2.5, 5 and 10 mg/kg, iv) produced a significant analgesic response in both post-partum and diestrous females using the hot water tail immersion latency test. However, the analgesic response in the post-partum females was significantly less than during diestrus at all doses tested. In addition, pretreatment with the mu 1 specific antagonist, Naloxonazine, significantly blunted the analgesic response in diestrous females, but did not significantly affect analgesia in post-partum females. These results indicate that morphine is less effective in producing analgesia in post-partum females. The mu 1 opiate receptor site does not appear to be involved in the analgesia produced during the post-partum period.

Multiple opiate receptor subtypes are involved in the stimulation of growth hormone release by beta-endorphin in female rats

The growth hormone (GH) secretory response to beta-endorphin and the involvement of opiate receptor subtypes in this response were determined in diestrous female rats. The involvement of the mu (mu), delta (delta) and/or kappa (kappa) site was determined by administering specific antagonists for each of these sites prior to beta-endorphin. beta-Funaltrexamine (1 or 5 micrograms) was administered to block mu sites, ICI 154,129 (5 or 25 micrograms) blocked delta sites and nor-binaltorphimine (8 micrograms) blocked kappa sites. The ability of these antagonists to block GH secretion following intravenous morphine administration was also determined. The opiate antagonists and beta-endorphin were administered into the lateral ventricle. A dose-response study for beta-endorphin indicated that 0.5 micrograms beta-endorphin was the minimum stimulatory dose for GH release, producing an approximately 4-fold increase in circulating levels of GH; lower doses of beta-endorphin did not stimulate secretion. All three antagonists were capable of blocking the stimulatory effects of beta-endorphin. These results provide evidence that all three opiate receptor subtypes are involved in the stimulatory effect of beta-endorphin on GH release.

Effects of age and gender on the AII-induced stimulation of prolactin release and inositol phosphate accumulation in rat anterior pituitary cells in vitro.

The stimulatory effects of Angiotensin II (AII) on prolactin secretion and inositol phosphate accumulation were examined in dispersed anterior pituitary cells collected from young (3-4 month), mature (7-8 month) and old (18-20 month) male and female rats. Physiological doses of AII (0.01-10 nM) stimulated prolactin release from cells collected from mature female rats only. This effect was antagonized by pretreatment with Saralasin, an AII receptor antagonist. Significant accumulation of the inositol phosphates was observed in cells obtained from the mature, female donors and this increase preceded the prolactin response. Although there was a small increase in total inositol phosphate accumulation in cells obtained from the old female rats, this was transient and did not coincide with a similar increase in prolactin release. These results indicate that pituitary sensitivity to AII stimulation is related to the age and the gender of the donor animal. The physiological role of pituitary AII needs to be examined in sexually mature female animals.