Arginine Vasopressin mRNA Levels Research Articles

Role of arginine vasopressin and corticotropin-releasing factor in mediating alcohol-induced adrenocorticotropin and vasopressin secretion in male rats bearing lesions of the paraventricular nuclei

In male rats, lesions of the paraventricular nucleus (PVN) of the hypothalamus attenuate, but do not abolish, adrenocorticotropin (ACTH) secretion in response to acute alcohol injection. As the PVN is the major source of corticotropin-releasing factor (CRF) in the median eminence, this observation suggests that extra-PVN brain regions, and/or ACTH secretagogues other than CRF (e.g. arginine vasopressin (AVP)), mediate ACTH stimulation by alcohol. This hypothesis was tested by examining the effect of AVP immunoneutralization in PVN-lesioned (PVNx) rats. Removal of endogenous AVP diminished alcohol-evoked ACTH secretion in both sham-operated and PVNx animals, indicating that AVP from outside the PVN partially mediates the hypothalamic-pituitary-adrenal (HPA) axis response to alcohol. This led us to determine whether alcohol might also regulate AVP steady-state gene expression in the supraoptic nucleus (SON) and PVN, and/or CRF mRNA in the PVN and the central nucleus of the amygdala (AMY). In the magnocellular portion of the PVN, sham-operated animals showed significantly increased PVN levels of both CRF and AVP mRNAs 3 h after alcohol. In the SON, alcohol administration tended to decrease AVP gene expression in PVNx rats, while the drug increased AVP mRNA levels in the SON of sham-operated rats. AMY levels of CRF mRNA were unaffected by these manipulations. Finally, since the regulation of alcohol-induced AVP mRNA levels in the SON appeared to depend on the presence of the PVN, we measured peripheral levels of AVP in both sham-operated and PVNx animals after injection of vehicle or alcohol. Although AVP decreased in all groups, alcohol depressed AVP secretion to a greater extent in PVNx animals, suggesting that AVP systems are more sensitive to inhibition in the absence of the PVN. Our results demonstrate that although AVP of PVN origin may participate in regulating the stimulatory effect to AVP on ACTH secretion, AVP from areas other than the PVN also plays a role. Additionally, regulation of both AVP gene expression in the SON and secretion in the systemic circulation are altered in rats bearing lesions of the PVN.

The neurosteroid tetrahydroprogesterone attenuates the endocrine response to stress and exerts glucocorticoid-like effects on vasopressin gene transcription in the rat hypothalamus

The neurosteroid tetrahydroprogesterone (5 alpha-pregnan-3 alpha-ol-20-one, allopregnanolone, THP), has been previously shown to counteract the anxiogenic effects of corticotropin-releasing hormone (CRH) and to interfere with noradrenergic and corticosteroid-mediated regulation of CRH release and gene transcription. Those observations indicated that, besides its sedative and analgesic activity, THP may also affect the neuroendocrine response to stress in a mode resembling that of corticosteroids. To examine this possibility, we compared the ability of THP, its precursor progesterone (P4), and the glucocorticoids dexamethasone (DEX) and corticosterone (CORT) to influence the pituitary-adrenal response to acute emotional stress and the adrenalectomy-induced increase in the gene transcription of the stress-related peptide arginine vasopressin (AVP) and of corticosteroid receptors (MR and GR) in the brain. Pretreatment of rats with a single dose of THP or P4 (50 micrograms/kg) significantly attenuated the elevation of plasma adrenocorticotropin (ACTH) and serum corticosterone after emotional stress; both steroids were, however, less potent than a similar dose of DEX. Administration of 1 mg of THP, CORT, or P4 to adrenalectomized (ADX) rats attenuated the increase in AVP mRNA levels in the ventromedial subdivision of the hypothalamic paraventricular nucleus (PVN), as compared with vehicle-treated ADX rats. However, whereas CORT and P4 influenced the ADX-induced increase in the transcription of both types of corticosteroid receptors in the hippocampus, these were unaffected by THP. In contrast to the glucocorticoids, THP and P4 failed to decrease plasma ACTH levels in rats deprived of endogenous steroids. These results demonstrate that the neurosteroid THP and its precursor P4 resemble glucocorticoids in their suppression of the pituitary-adrenal response to emotional stress; however, THP influences the transcription of glucocorticoid-responsive genes in brain structures involved in the regulation of the hypothalamo-pituitary-adrenal system in a fashion that is quite distinct from that obtained with glucocorticoids.

Differential regulation of oxytocin and vasopressin messenger ribonucleic acid levels by gonadal steroids in postpartum rats

We previously reported that sequential estradiol and progesterone exposure followed by progesterone withdrawal increases oxytocin (OT), but not arginine vasopressin (AVP), messenger ribonucleic acid (mRNA) in the hypothalamic paraventricular nucleus (PVN) of the rat. Substitution of testosterone for progesterone and subsequent testosterone withdrawal in the estrogen-primed rat increases PVN AVP mRNA levels. At the end of pregnancy (day 21), rats are exposed to high estrogen and declining progesterone and testosterone concentrations. Coincident with these changes in circulating gonadal steroid hormones are increases in OT and AVP mRNAs. If progesterone levels are sustained at term, OT levels are attenuated and if testosterone is sustained, AVP mRNA levels are attenuated. Immediately postpartum, however, OT and AVP mRNA levels decline compared to term levels. To further determine the role of estrogen in the regulation of OT and AVP mRNAs, we performed two experiments. In the first experiment, we administered estrogen during the peripartum period to determine if estrogen supplementation prevents the relative attenuation of OT and AVP mRNAs that is seen after parturition. Day 18 pregnant rats were given estradiol-filled or empty capsules and sacrificed on day 2 of lactation. By Northern analysis, significant differences in PVN AVP, but not OT, mRNA were found between the estrogen- and sham-treated lactational animals, P < 0.02. In the second experiment, we determined if sustaining estrogen after progesterone is removed in steroid-treated ovariectomized rats is essential for the increase in OT mRNA. Ovariectomized rats were given either empty capsules or sequential estradiol- and progesterone-filled capsules and both were sustained for 12 days. When progesterone-filled capsules were removed, estradiol-filled capsules were either removed or left in place, and the animals were sacrificed 48 h later. PVN OT mRNA was analyzed by Northern blot hybridization. OT mRNA was increased in both of the steroid-treated groups to the same degree, compared to sham-treated animals, P = 0.04. In summary, estrogen supplementation during early lactation prevents the attenuation of PVN AVP, but not OT, mRNA after parturition. In the estrogen-primed ovariectomized rat, it is not necessary to sustain estrogen to see the effects of progesterone withdrawal upon PVN OT mRNA.

Sequential exposure to estrogen and testosterone (T) and subsequent withdrawal of T increases the level of arginine vasopressin messenger ribonucleic acid in the hypothalamic paraventricular nucleus of the female rat.

The hypothalamic peptides arginine vasopressin (AVP) and oxytocin (OT) have been implicated as mediators of socio-sexual behaviors in addition to their roles in osmolar homeostasis (AVP), milk ejection and uterine contractility (OT). Within 24 h of parturition, OT and AVP messenger ribonucleic acid (mRNA) levels increase in the hypothalamic paraventricular, and to a lesser degree, the supraoptic nucleus (PVN and SON) of the rat. We previously reported that the prepartum increase in OT mRNA is related to the spontaneous decline in progesterone levels prior to parturition. We also reported that increases in PVN and SON OT mRNA can be induced by exposing the ovariectomized rat to a steroid regimen that mimics the steroid milieu of pregnancy, namely sequential estrogen and progesterone and subsequent progesterone withdrawal. Levels of PVN and SON AVP mRNAs were not affected by progesterone withdrawal in late pregnant rats or the steroid regimen that increased OT mRNA in ovariectomized rats. These observations suggest that other factors, perhaps hormonal, may influence AVP mRNA levels. A decline in testosterone coincident with waning progesterone levels also occurs prepartum. Since peak levels of AVP mRNA prepartum coincide with the prepartum decline in testosterone, we questioned whether declining testosterone levels are important for the increase in AVP mRNA levels. To examine a possible role for testosterone in the increased level of AVP mRNA in late pregnancy, we sequentially administered estradiol and testosterone long-term (2 weeks) and removed testosterone 48 h prior to sacrifice. This steroid regimen mimics the estrogen and testosterone pattern of late pregnancy in rats. AVP, but not OT, mRNA levels increased significantly in the PVN of ovariectomized rats receiving this steroid regimen. We also found that implantation of late pregnant rats with testosterone capsules to prevent the spontaneous prepartum decline in testosterone, attenuates the increase in PVN AVP, but not OT, mRNA on day 21 of pregnancy. The data show that sequential estrogen and testosterone and testosterone withdrawal increase the level of PVN AVP mRNA in the female rat.

Osmotic and non-osmotic regulation of arginine vasopressin (AVP) release, mRNA, and promoter activity in small cell lung carcinoma (SCLC) cells

Arginine vasopressin (AVP) is synthesized in the magnocellular neurons of the hypothalamus and stored in the posterior pituitary. It has been shown that hypothalamic AVP mRNA is increased during experimental stimulation of osmotic and non-osmotic stimulation of AVP release. The mechanisms underlying the stimulation of AVP biosynthesis in these conditions are not known. The present study was, therefore, performed to measure AVP release, AVP mRNA level, and AVP gene promoter activity during osmotic and non-osmotic stimulation of AVP secretion in the small cell lung carcinoma (SCLC) cells. AVP release was measured by radioimmunoassay, steady state levels of AVP mRNA by solution hybridization, and AVP gene promoter activity exhibited by a 1.5 kb 5′-flanking AVP gene fragment fused to a luciferase reporter after SCLC cells were subjected to osmotic or non-osmotic conditions. High media osmolality (330 mOsm) significantly increased AVP release (control (C) 1.42 ± 0.27 vs. High Osm 3.67 ± 0.39 pg/2 × 10 6 cells, N = 9, P < 0.002); AVP mRNA (C 173.6 ± 16.8 vs. High Osm 280.1 ± 19.4 pg/2 × 10 6 cells, N = 7, P < 0.001); and AVP gene promoter activity (C 1353 ± 99 vs. High Osm 2026 ± 134 L.U./10 −4 U β-gal, N = 8, P < 0.001). Non-osmotic stimulators, 0.1 μM endothelin 3 (ET3), 1 μM angiotensin II (AII), and 10 μM acetylcholine (Ach) significantly increased AVP release; ET3 (C 1.78 ± 0.20 vs. ET3 6.85 ± 1.86 pg/2 × 10 6 cells, N = 8, P < 0.02); AII (C 1.29 ± 0.38 vs. AII 27.80 ± 7.09 pg/2 × 10 6 cells, N = 5, P < 0.05) and Ach (C 1.14 ± 0.33 vs. Ach 2.68 ± 0.58 pg/2 × x10 6 cells, N = 6, P < 0.05). However, only ET3 significantly increased AVP mRNA (C 166.6 ± 19.6 vs. ET3 254.4 ± 25.6 pg/p × 10 6 cells, N = 5, P < 0.05) and AVP promoter activity (C 1515 ± 163 vs. ET3 2389 ± 342 L.U./10 −4 U β-gal, n = 6, P < 0.05). To localize the region of the AVP promoter that mediates the osmotic stimulation and the effect of ET3, 5′ deletions of the AVP promoter fragments terminating at −532, −211, and −102, was assessed. Only the promoter activity of the 1.5 kb construct, but not the delection constructs, was significantly increased by ET3 or high osmolality. These results suggest that modulation of AVP gene transcription is, at least in part, responsible for increased AVP synthesis and release in response to osmotic and non-osmotic stimulation, and that the region of 5′ flanking sequence between −1500 and −532 contains the elements responsible for the effects.

Increased expression of corticotropin-releasing hormone and vasopressin messenger ribonucleic acid (mRNA) in the hypothalamic paraventricular nucleus during repeated stress: association with reduction in glucocorticoid receptor mRNA levels.

Hypothalamic-pituitary-adrenal (HPA) responses remain intact or increase after chronic or repeated stress despite robust levels of circulating glucocorticoids that would be expected to restrain the responsiveness of the axis. The purpose of this study was to determine whether chronic stress altered corticosteroid receptor messenger RNA (mRNA) levels at any locus known to mediate glucocorticoid feedback on HPA function (i.e. hippocampus or hypothalamus), whether such effects were glucocorticoid dependent, and whether changes in corticosteroid receptor function could potentially contribute to the putative shift from corticotropin-releasing hormone (CRH) to arginine vasopressin (AVP) in the hypothalamic paraventricular nucleus (PVN) in the modulation of pituitary adrenal function occurring during chronic stress. We compared the stress responsiveness of sham-operated rats to that of adrenalectomized rats using a moderate dose of corticosterone (CORT) pellet replacement (ADX + CORT group). Acute immobilization caused a significant increase in CRH, but not AVP, mRNA levels in the parvocellular PVN in sham rats. The ADX + CORT group showed significantly greater increases in both CRH and AVP mRNA levels in the PVN compared to sham rats. These data indicate that PVN AVP mRNA levels are more sensitive to glucocorticoid negative feedback than are the levels of CRH mRNA. In repeated stress, the sham groups showed robust increases in PVN CRH and AVP mRNA levels despite high levels of plasma CORT. The rise in AVP mRNA levels was greater than that in CRH mRNA. Type II glucocorticoid receptor mRNA in the hippocampus and PVN was decreased in the repeatedly stressed sham group. These data suggest a decrease in the CORT negative feedback restraint of PVN CRH and AVP mRNA levels repeated stress and a persistence of relatively greater responsiveness of AVP mRNA levels to CORT negative feedback. After repeated stress in ADX+CORT rats, both PVN CRH and AVP mRNA levels showed robust responses, with a relatively greater increase in AVP mRNA. These data indicate that a CORT-mediated decrease in hippocampal and hypothalamic glucocorticoid receptor mRNA levels is not the only mechanism contributing to the maintenance of a robust HPA response after repeated stress. Similarly, we postulate that the relative shift from CRH to AVP in the PVN after repeated stress is mediated by both a greater sensitivity of AVP to CORT negative feedback and CORT-independent mechanisms.

Thyroidectomy does not alter hypothalamic oxytocin and vasopressin expression in chronically hypernatremic rats

Sustained hyperosmolality increases the levels of hypothalamic oxytocin (OT) and arginine vasopressin (AVP) messenger ribonucleic acids (mRNAs). Gonadectomy is known to abolish this response (12,18). In this study we investigated whether thyroidectomy would alter OT and AVP mRNA levels in the hypothalamic paraventricular nucleus (PVN) of the hyperosmotically stimulated rat. Male Sprague-Dawley rats underwent thyroidectomy (hypothyroid) or sham thyroidectomy (euthyroid) at 7 weeks of age. Three weeks later hypothyroid and euthyroid animals were administered 2% NaCl (6–11 days) or tap water and sacrificed at the end of the experiment. Northern blot hybridization was used to assess size and levels of hypothalamic OT and AVP mRNAs. Hypothyroid rats had significantly lower levels of serum thyroxine (T 4) than their euthyroid cohorts ( P < 0.0001). Both the euthyroid and the hypothyroid animals receiving 2% NaCl developed hypernatremia and increased the levels and the size of OT and AVP mRNAs compared to their tap water cohorts. We conclude that in contrast to gonadectomy, thyroidectomy does not alter the level of OT and AVP mRNAs in the hypothalamus of chronically hypernatremic male rats.

Dehydration stimulates hypothalamic gene expression of histamine synthesis enzyme: importance for neuroendocrine regulation of vasopressin and oxytocin secretion

Dehydration associated with hyperosmolality and decreased extracellular volume stimulates arginine vasopressin (AVP) and oxytocin (OT) secretion from magnocellular neurons of the hypothalamus. The effects of hyperosmolality and decreased extracellular volume on the magnocellular neurons are mainly indirect and seem to be mediated centrally via several neurotransmitters and neuropeptides. Because histamine (HA), which serves as a central neurotransmitter, releases AVP and OT from the neurohypophysis when administered centrally, we investigated the possible role of HA in dehydration-induced AVP and OT secretion. To do this, we studied 1) the effect of dehydration on messenger RNA (mRNA) expression of the HA synthesis enzyme, histidine decarboxylase (HDC), in the tuberomammillary nucleus of the hypothalamus; and 2) the effect of HA synthesis inhibition during dehydration on AVP and OT mRNA expression in the supraoptic nucleus of the hypothalamus as well as on plasma AVP and OT levels. Forty-eight hours of dehydration increased the mRNA level of HDC in the tuberomammillary nuclei, whereas 24 h of dehydration had no effect. Pretreatment with the HA synthesis inhibitor alpha-fluoromethylhistidine (alpha FMH) increased the expression of HDC mRNA in 24-h dehydrated rats, but had no effect in euhydrated rats. In rats dehydrated for 48 h, the already increased level of HDC mRNA was not increased further by alpha FMH. Twenty-four and 48 h of dehydration increased AVP and OT mRNA levels in the supraoptic nucleus. This effect was inhibited by alpha FMH pretreatment. Dehydration increased the plasma levels of AVP and OT to an extent which depended on the duration of dehydration. Pretreatment with alpha FMH inhibited the hormone responses to 24 h of dehydration, but did not affect the responses to 48 h of dehydration. Twenty-four and 48 h of dehydration had no significant effect on the contents of AVP and OT in the neurohypophysis, whereas pretreatment with alpha FMH combined with 48 h of dehydration led to depletion of AVP stores in the neurohypophysis. Based on the present findings, we conclude that hypothalamic histaminergic neurons are involved in the regulation of dehydration-induced stimulation of magnocellular AVP and OT neurons.

Dehydration stimulates hypothalamic gene expression of histamine synthesis enzyme: importance for neuroendocrine regulation of vasopressin and oxytocin secretion.

Dehydration associated with hyperosmolality and decreased extracellular volume stimulates arginine vasopressin (AVP) and oxytocin (OT) secretion from magnocellular neurons of the hypothalamus. The effects of hyperosmolality and decreased extracellular volume on the magnocellular neurons are mainly indirect and seem to be mediated centrally via several neurotransmitters and neuropeptides. Because histamine (HA), which serves as a central neurotransmitter, releases AVP and OT from the neurohypophysis when administered centrally, we investigated the possible role of HA in dehydration-induced AVP and OT secretion. To do this, we studied 1) the effect of dehydration on messenger RNA (mRNA) expression of the HA synthesis enzyme, histidine decarboxylase (HDC), in the tuberomammillary nucleus of the hypothalamus; and 2) the effect of HA synthesis inhibition during dehydration on AVP and OT mRNA expression in the supraoptic nucleus of the hypothalamus as well as on plasma AVP and OT levels. Forty-eight hours of dehydration increased the mRNA level of HDC in the tuberomammillary nuclei, whereas 24 h of dehydration had no effect. Pretreatment with the HA synthesis inhibitor alpha-fluoromethylhistidine (alpha FMH) increased the expression of HDC mRNA in 24-h dehydrated rats, but had no effect in euhydrated rats. In rats dehydrated for 48 h, the already increased level of HDC mRNA was not increased further by alpha FMH. Twenty-four and 48 h of dehydration increased AVP and OT mRNA levels in the supraoptic nucleus. This effect was inhibited by alpha FMH pretreatment. Dehydration increased the plasma levels of AVP and OT to an extent which depended on the duration of dehydration. Pretreatment with alpha FMH inhibited the hormone responses to 24 h of dehydration, but did not affect the responses to 48 h of dehydration. Twenty-four and 48 h of dehydration had no significant effect on the contents of AVP and OT in the neurohypophysis, whereas pretreatment with alpha FMH combined with 48 h of dehydration led to depletion of AVP stores in the neurohypophysis. Based on the present findings, we conclude that hypothalamic histaminergic neurons are involved in the regulation of dehydration-induced stimulation of magnocellular AVP and OT neurons.

Effect of testosterone and its metabolites upon the level of vasopressin messenger ribonucleic acid in the hypothalamus of the hyperosmotically stimulated male rat.

We previously reported that gonadal steroids modify the expression of arginine vasopressin (AVP) in the hypothalamus of rats administered 2% sodium chloride solution for 5 days. Gonadectomy prevented, and testosterone (T) replacement restored, enhanced AVP mRNA levels in the supraoptic nucleus (SON) of male rats receiving this hyperosmotic challenge. The present study investigated the effects of the androgenic and estrogenic metabolites of T on hypothalamic AVP mRNA levels in response to chronic hyperosmolality. Gonadectomized male rats receiving 2% NaCl for 5 days and treated with T, dihydrotestosterone (DHT), or DHT+estradiol (E2), but not E2 alone or empty implants, had increased AVP mRNA levels compared to gonadectomized animals receiving tap water. Our results support a role for T and DHT-mediated effects upon the enhanced accumulation of AVP mRNA in the SON of male rats receiving a chronic hyperosmotic challenge.

Effects of adrenalectomy and Type I or Type II glucocorticoid receptor activation on AVP and CRH mRNA in the rat hypothalamus

The brain contains two types of adrenal steroid receptors, which play a role in mediating adrenal steroid effects on neuropeptide and other types of gene expression in discrete brain regions. Because the paraventricular nuclei of the hypothalamus (PVN) have adrenal steroid-sensitive neuropeptide systems, they provide a bench-mark to assess the doses of receptor agonists that may act selectively via Type I and Type II receptors. In the present study, in situ hybridization histochemistry was used to examine the effects of adrenalectomy (ADX) and Type I and Type II receptor agonists on arginine vasopressin (AVP) mRNA and corticotropin-releasing hormone (CRH) mRNA in rat brain. In agreement with previous reports, adrenal steroid regulation of AVP and CRH mRNA was found to be mediated primarily through the Type II receptor. Furthermore, adrenalectomy significantly increased AVP mRNA in the parvocelluar region of the hypothalamic paraventricular nucleus (PVN), and systemic administration of the specific Type II agonist, RU28362 (10 μg/μ1/h), as well as corticosterone (CORT) pellets of 50 and 300 mg, prevented this increase. CRH mRNA was not significantly increased after ADX, but was markedly decreased in the PVN of rats receiving either RU28362 or a 300 mg pellet of CORT. Aldosterone, a specific Type I agonist, did not significantly affect either AVP or CRH mRNA levels when administered at 10 μg/h. Moreover, in the magnocellular regions of the PVN and SON AVP mRNA did not vary as a function of steroid manipulation. Results for PVN are compared to changes in gene products in hippocampus produced by the same doses of Type I and Type II agonists, in which Type I receptors appear to have a significant role.

Circadian variation of arginine-vasopressin messenger RNA in the rat suprachiasmatic nucleus

Arginine-vasopressin (AVP) gene expression in the rat suprachiasmatic nucleus (SCN) is subject to daily rhythmic changes. To determine whether this variation is endogenously generated, temporal changes in the SCN AVP mRNA level in constant dark (DD) condition was compared with changes occurring under the light-dark (LD) condition. In both lighting conditions, the presence of a rhythm in AVP mRNA level was observed in the SCN. In LD condition, peak level of AVP mRNA was found during the latter part of the day (zeitgeber time or ZT 8) and trough value during the night at ZT 20. Correspondingly, peak level of AVP mRNA under DD condition was observed during the latter part of the subjective day (circadian time or CT 8) and a trough during the subjective night (CT 20). Under both lighting conditions, a rapid increase and decrease of mRNA around the peak time was also observed. On the other hand, no significant daily variation in AVP mRNA was found in the supraoptic nucleus in both LD and DD conditions. These results provide evidence that a rhythmic change in AVP mRNA level is regulated by a circadian clock intrinsic to the SCN. The phase relationship of AVP mRNA rhythm to peptide rhythm in the SCN is discussed.

Male Fischer 344/N rats show a progressive central impairment of the hypothalamic-pituitary-adrenal axis with advancing age

We investigated the effects of aging on the regulation of hypothalamic-pituitary-adrenal function and hippocampal steroid receptors in a series of in vivo and in vitro studies conducted in healthy intact 2-, 8-, 18-, and 24-month-old male Fischer 344/N rats. Basal plasma ACTH levels were similar among age groups, and basal plasma corticosterone levels showed a significant aging-associated decline. Two i.v. doses (2 and 20 micrograms/kg BW) of rat CRF elicited significantly greater and delayed ACTH and greater corticosterone responses in older rats, consistent with the pattern encountered in hypothalamic CRF deficiency. In contrast, the i.v. injection of a muscarinic agonist, arecoline, elicited similar ACTH and corticosterone responses in all age groups. An i.v. injection of ACTH-(1-24) evoked lower corticosterone responses in the older (18- and 24-month-old) than in the younger (2- and 8-month-old) groups of rats, consistent with an impairment of hypothalamic-pituitary-adrenal axis function in older animals. Steady state mRNA levels of mineralocorticoid and glucocorticoid receptors were significantly decreased in the hippocampus of the 8-, 18-, and 24-month-old rats, compatible with maturational, rather than senescent, changes. CRF mRNA levels in the paraventricular nucleus of the hypothalamus, CRF content, and in vitro secretion by whole explanted hypothalami were progressively and significantly reduced with age, whereas the steady state levels of arginine vasopressin mRNA were significantly increased with age. Steady state levels of POMC mRNA were decreased, and ACTH content and in vitro secretion by corticotrophs were increased with age in the anterior pituitary. We conclude that male Fischer 344/N rats show a progressive hypothalamic CRH deficiency with advancing age, which appears to be associated with elevated production of arginine vasopressin in the hypothalamus.

Male Fischer 344/N rats show a progressive central impairment of the hypothalamic-pituitary-adrenal axis with advancing age.

We investigated the effects of aging on the regulation of hypothalamic-pituitary-adrenal function and hippocampal steroid receptors in a series of in vivo and in vitro studies conducted in healthy intact 2-, 8-, 18-, and 24-month-old male Fischer 344/N rats. Basal plasma ACTH levels were similar among age groups, and basal plasma corticosterone levels showed a significant aging-associated decline. Two i.v. doses (2 and 20 micrograms/kg BW) of rat CRF elicited significantly greater and delayed ACTH and greater corticosterone responses in older rats, consistent with the pattern encountered in hypothalamic CRF deficiency. In contrast, the i.v. injection of a muscarinic agonist, arecoline, elicited similar ACTH and corticosterone responses in all age groups. An i.v. injection of ACTH-(1-24) evoked lower corticosterone responses in the older (18- and 24-month-old) than in the younger (2- and 8-month-old) groups of rats, consistent with an impairment of hypothalamic-pituitary-adrenal axis function in older animals. Steady state mRNA levels of mineralocorticoid and glucocorticoid receptors were significantly decreased in the hippocampus of the 8-, 18-, and 24-month-old rats, compatible with maturational, rather than senescent, changes. CRF mRNA levels in the paraventricular nucleus of the hypothalamus, CRF content, and in vitro secretion by whole explanted hypothalami were progressively and significantly reduced with age, whereas the steady state levels of arginine vasopressin mRNA were significantly increased with age. Steady state levels of POMC mRNA were decreased, and ACTH content and in vitro secretion by corticotrophs were increased with age in the anterior pituitary. We conclude that male Fischer 344/N rats show a progressive hypothalamic CRH deficiency with advancing age, which appears to be associated with elevated production of arginine vasopressin in the hypothalamus.

The effect of salt-loading on corticotropin releasing hormone and arginine vasopressin mRNA levels in the mouse hypothalamus: A quantitative in situ hybridization analysis

Previously, we showed that during salt-loading in mice there was an acute rise in plasma ACTH levels after 2 days followed by a transient decrease after 4 and 9 days. Pro-opiomelanocortin (POMC) mRNA levels in the anterior pituitary increased after 2 days and returned to normal thereafter. In this study, changes in hypothalamic CRH and AVP mRNA levels during salt-loading were investigated using quantitative in situ hybridization histochemistry. CRH mRNA was expressed only in the paraventricular nucleus (PVN), while AVP mRNA was expressed in both the supraoptic (SON) and paraventricular nuclei. CRH mRNA levels were unchanged after 2 days salt-loading, but declined to 77% of control levels after 9 days. AVP mRNA levels rose to 260% and 634% of control levels in the SON, and to 352% and 522% of control levels in the PVN, after 2 and 9 days salt-loading, respectively. These data suggest a major role of AVP in the acute stimulation of ACTH secretion and POMC mRNA levels seen after 2 days salt-loading. Densitization of AVP receptors at the corticotroph level and a centrally mediated inhibition of CRH release may account for the decrease of ACTH secretion and POMC mRNA levels in the anterior pituitary with prolonged salt-loading.

Local tetrodotoxin blocks chronic stress effects on corticotropin-releasing factor and vasopressin messenger ribonucleic acids in hypophysiotropic neurons.

To test the hypothesis that synaptic inputs to the paraventricular nucleus mediate stress-induced increases in corticotropin-releasing peptide expression in the paraventricular nucleus of the hypothalamus (PVH), relative levels of the mRNAs encoding corticotropin-releasing factor (CRF) and arginine vasopressin (AVP) were followed, in situ, in animals subjected to chronic footshock stress and concurrent local administration of tetrodotoxin or vehicle. Consistent with previous findings, a 7-day exposure to chronic footshock resulted in a 2.1-fold increase in CRF mRNA levels in the parvocellular division of the PVH. The footshock paradigm also resulted in at least a 41% increase in AVP transcripts in this same region; this effect was localized predominantly to CRF-immunoreactive neurons. The stressor did not significantly alter AVP mRNA levels in the magnocellular division of the PVH. Tetrodotoxin, administered to the PVH via osmotic minipump, blocked the stress-induced rise in CRF and AVP mRNAs, but had no significant effect on basal levels of these transcripts. The results support the view that maintenance of the enhanced central drive on pituitary-adrenal activity seen in response to chronic stress is mediated via neural inputs to the PVH.

Selective forebrain fiber tract lesions implicate ventral hippocampal structures in tonic regulation of paraventricular nucleus corticotropin-releasing hormone (CRH) and arginine vasopressin (AVP) mRNA expression

The hippocampus appears to be involved in tonic regulation of the hypothalamo-pituitary-adrenocortical axis via interactions with corticotropin-releasing hormone (CRH) and arginine vasopressin (AVP)-containing neurons of the hypothalamic paraventricular nucleus (PVN). To further investigate the anatomical basis of such interactions, lesions were made to forebrain fiber tracts in position to communicate inhibitory information from the hippocampus to the PVN. Total fimbria-fornix transections (TFF) and lateral fimbria-fornix lesions (LFF) both significantly increased CRH mRNA levels in the medial parvocellular PVN, as assayed by semi-quantitative in situ hybridization histochemistry. Medial fimbria-fornix lesions or section of the medial corticohypothalamic tracts (MCHT) did not influence CRH mRNA levels. The LFF group showed increases in both AVP mRNA and ACTH secretion, whereas no other lesion was effective in this regard. The results suggest: (1) hippocampal efferents confering tonic inhibition of the HPA axis probably originate in regions contributing to the lateral extent of the fornix, representing structures in the ventral subiculum and ventral extent of CA1: (2) projections from the hippocampus to the medial basal hypothalamus (travelling in the MCHT) are unlikely to affect HPA function: (3) hippocampus may influence the PVN CRH/AVP neuron at multiple levels, in that LFF and TFF lesions have differential effects on PVN AVP mRNA levels and ACTH secretion.

The magnocellular arginine-vasopressin mRNA responds differently to food deprivation between the supraoptic and paraventricular nuclei of the hypothalamus in adrenalectomized rats with low corticosterone replacement

We previously reported that food deprivation significantly decreased arginine-vasopressin (AVP) mRNA levels in the supraoptic (SON) and paraventricular (PVN) nuclei of the hypothalamus and also greatly stimulated the pituitary-adrenocortical system in rats. In this study, we deprived adrenalectomized rats with subcutaneously implanted low-dose corticosterone pellets (ADX + B) of food for 3 days to investigate the involvement of corticosteroid feedback regulation in the food deprivation-induced decrease in AVP mRNA in both the SON and the PVN. The plasma corticosterone levels in these animals were maintained at low levels constantly over 24 h. The ACTH concentration in the morning plasma was markedly increased in the food-deprived ADX + B rats as compared to the fed ADX + B rats. Food deprivation significantly decreased the corticotropin-releasing hormone (CRH) content in the median eminence and increased the CRH and AVP content in the neurointermediate lobe of the pituitary. Semiquantitative in situ hybridization histochemistry revealed that AVP mRNA levels were decreased in the SON but, inversely, increased in magnocellular as well as parvocellular subdivisions of the PVN following food deprivation. These results suggest that: (1) AVP mRNA responds differently to food deprivation between the SON and the PVN; (2) the glucocorticoid feedback can exert on AVP mRNA in the PVN but not in the SON in the food-deprived rats; and (3) food deprivation affects the neurohypophysial levels of CRH and AVP.

Hypothalamic-pituitary-adrenal axis function in the Zucker obese rat.

The development of many endocrine, metabolic, and behavioral abnormalities characteristic of genetically obese Zucker rats is dependent upon the presence of glucocorticoids, the secretion of which is regulated by a neuroendocrine cascade initiated by hypothalamic release of CRF. Recent reports have inferred alterations in central CRF tone as a putative factor contributing to dysregulation of the pituitary-adrenal axis and of metabolic processes in this phenotype. In the current study the hypothalamic CRF system in Zucker lean (FA/?) and obese (fa/fa) phenotypes was functionally evaluated. Neither the stalk median eminence content of CRF or arginine vasopressin (AVP) nor hypothalamic levels of CRF or AVP mRNA differed in the lean and obese phenotypes. No phenotypic differences were observed in either basal or stimulated CRF release from hypothalamic tissue obtained from lean and obese rats. Furthermore, in intact rats the magnitude of pituitary-adrenal responses to various stressors was also similar between phenotypes. However, secretion of CRF and AVP into the hypophysial-portal circulation of obese rats was, respectively, 73% and 35% lower than that of the lean rats. Adrenalectomy was associated with a 3-fold elevation of hypophysial-portal CRF levels in obese rats compared to intact controls. Corticosterone infusion was more effective in suppressing portal CRF levels in adrenalectomized obese compared to adrenalectomized lean rats. Finally, neither CRF receptor number and affinity nor the magnitude of pituitary-adrenal responses to rat CRF challenge (1 micrograms, iv) differed between Zucker phenotypes. These observations lead us to infer that rats of the obese phenotype exhibit reduced hypothalamic CRF tone due to dysregulation of the HPA axis at a site proximal to the hypophysiotropic CRF system that mediates glucocorticoid feedback regulation.

Ontogeny of Expression and Glucocorticoid Regulation of the Arginine Vasopressin Gene in the Rat Hypothalamic Paraventricular Nucleus

Corticotropin-releasing factor and arginine vasopressin (AVP) are the two major hypothalamic factors that regulate anterior pituitary adrenocorticotropin secretion during stress. We have previously reported that the expression of the corticotröpin-releasing factor gene in the hypothalamus and its regulation by glucocorticoids were not mature during the first week of life in the rat, i.e. during the stress non-responsive period. In this report, we studied the ontogeny of expression of the AVP gene in the hypothalamic paraventricular nucleus in rats using in situ hybridization. AVP mRNA was detected as early as day 20 of gestation (E20) both in the parvocellular and the magnocellular portion of the paraventricular nucleus. The levels of expression of the AVP gene increased steadily from E20 to the third day after birth (P3) and remained stable from P3 to P14. Bilateral surgical adrenalectomy induced an increase in AVP mRNA levels in the parvocellular portion of the paraventricular nucleus, but not in the magnocellular portion, in both 7-day-old and 14-day-old rats, suggesting that the glucocorticoid regulation of the AVP synthesizing neurons of the paraventricular nucleus is mature in the developing postnatal rat.