Changes in plasma arginine vasopressin concentrations in cyclists participating in a 109-km cycle race.

OBJECTIVE: To evaluate the osmotic and non-osmotic regulation of arginine vasopressin (AVP) during endurance cycling. DESIGN: Observational study. Setting 109 km cycle race. PARTICIPANTS: 33 Cyclists. INTERVENTIONS: None. MAIN OUTCOME MEASUREMENTS: Plasma sodium concentration ([Na(+)]), plasma volume (PV) and plasma arginine vasopressin (AVP) concentration ([AVP](p)). RESULTS: A fourfold increase in [AVP](p) occurred despite a 2-mmol l(-1) decrease in plasma [Na(+)] combined with only modest (5%) PV contraction. A significant inverse correlation was noted between [AVP](p) Delta and urine osmolality Delta (r = -0.41, p<0.05), whereas non-significant inverse correlations were noted between [AVP](p) and both plasma [Na(+)] Delta and % PV Delta. Four cyclists finished the race with asymptomatic hyponatraemia. The only significant difference between the entire cohort with this subset of athletes was postrace plasma [Na(+)] (137.7 vs 133.5 mmol l(-1), p<0.001) and plasma [Na(+)] Delta (-1.9 vs -5.1 mmol l(-1), p<0.05). The mean prerace [AVP](p) of these four cyclists was just below the minimum detectable limit (0.3 pg ml(-1)) and increased marginally (0.4 pg ml(-1)) despite the decline in plasma [Na(+)]. CONCLUSIONS: The osmotic regulation of [AVP](p) during competitive cycling was overshadowed by non-osmotic AVP secretion. The modest decrease in PV was not the primary non-osmotic stimulus to AVP. Partial suppression of AVP occurred in four (12%) cyclists who developed hyponatraemia during 5 h of riding. Therefore, these results confirm that non-osmotic AVP secretion and exercise-associated hyponatraemia does, in fact, occur in cyclists participating in a 109 km cycle race. However, the stimuli to AVP is likely different between cycling and running.

Acute changes in arginine vasopressin, sweat, urine and serum sodium concentrations in exercising humans: does a coordinated homeostatic relationship exist?

The parallel response of sweat rate and urine production to changes in plasma osmolality and volume support a role for arginine vasopressin (AVP) as the main endocrine regulator of both excretions. A maximal test to exhaustion and a steady-state run on a motorised treadmill were both completed by 10 moderately trained runners, 1 week apart. Sweat, urine and serum sodium concentrations ([Na+]) were evaluated in association with the plasma concentrations of cytokines, neurohypophyseal and natriuretic peptides, and adrenal steroid hormones. When data from both the high-intensity and steady-state runs were combined, significant linear correlations were noted between: sweat [Na+] versus postexercise urine [Na+] (r=0.80; p<0.001), postexercise serum [Na+] versus both postexercise urine [Na+] (r=0.56; p<0.05) and sweat [Na+] (r=0.64; p<0.01) and postexercise urine [Na+] versus postexercise plasma arginine vasopressin concentration ([AVP](P)) (r=0.48; p<0.05). A significant positive correlation was noted between postexercise [AVP](P) and sweat [Na+] during the steady-state condition only (r=0.66; p<0.05). These correlations suggest that changes in serum [Na+] during exercise may evoke corresponding changes in sweat and urine [Na+], which are likely regulated coordinately by changes in [AVP](P) to preserve body fluid homeostasis.

Oxytocin secretion in response to cholecystokinin and food: differentiation of nausea from satiety.

Administration of cholecystokinin (CCK) to rats caused a dose-dependent increase in plasma levels of the neurohypophyseal hormone oxytocin (OT). The OT secretion was comparable to that found in response to nausea-producing chemical agents that cause learned taste aversions. The effect of CCK on OT secretion was blunted after gastric vagotomy, as was the inhibition of food intake induced by CCK. Food ingestion also led to elevated plasma OT in rats, but CCK and aversive agents caused even greater OT stimulation. Thus, after administration of large doses of CCK, vagally mediated activation of central nausea pathways seems to be predominantly responsible for the subsequent decrease in food intake. Despite their dissimilar affective states, both nausea and satiety may activate a common hypothalamic oxytocinergic pathway that controls the inhibition of ingestion.

Vasopressin stimulation of adrenocorticotropin hormone (ACTH) in humans. In vivo bioassay of corticotropin-releasing factor (CRF) which provides evidence for CRF mediation of the diurnal rhythm of ACTH.

The diurnal response of ACTH release to intravenously administered arginine vasopressin was tested in normal volunteers given consecutively moderate doses of vasopressin every 15 min (0.1, 0.3, 1.0, and 3.0 IU) at 2200 h and again at 0700 h (PM/AM). This protocol was repeated 4 wk later with the times reversed (AM/PM). A dose-related increase in ACTH secretion was observed in all subjects. When the AM response of the AM/PM protocol was compared with the PM response of the PM/AM protocol, the release of ACTH was greater in the morning (P less than 0.05) as evaluated by the following criteria: peak value of ACTH (129.9 +/- 30.4 pg/ml in the AM vs. 57.1 +/- 20.2 in the PM); area under the curve (689 in the AM vs. 259 in the PM); and, sensitivity of the ACTH dose-response curve (first significant increase in ACTH with 1 IU of vasopressin in the AM but not significant even after 3 IU in the PM). In addition, when the AM vasopressin testing followed a previous evening stimulation (PM/AM protocol), there was a blunted ACTH response compared with the AM/PM protocol. Corticotropin-releasing factor (CRF) is probably the major ACTH secretagogue, but since vasopressin acts synergistically with CRF to produce an augmented release of ACTH, we suggest that the ACTH response to administered vasopressin depends upon the ambient endogenous level of CRF. We interpret our data and published data that CRF produces a lesser release of ACTH in the AM as follows: in the morning endogenous CRF is high and administered CRF produces little further release of ACTH, but administered vasopressin acting synergistically with high endogenous CRF causes a greater release of ACTH; conversely, in the evening endogenous CRF is low and administered CRF causes a greater release of ACTH, but vasopressin (a weak secretagogue by itself) gives a low ACTH response. We conclude that vasopressin stimulation of ACTH secretion can be used as an in vivo bioassay of endogenous CRF, and that there is a diurnal rhythm of CRF in hypophyseal portal blood.

Role of renal aquaporins in escape from vasopressin-induced antidiuresis in rat.

The purpose of this study was to investigate whether escape from vasopressin-induced antidiuresis is associated with altered regulation of any of the known aquaporin water channels. After 4-d pretreatment with 1-deamino-[8-D-arginine]-vasopressin (dDAVP) by osmotic mini-pump, rats were divided into two groups: control (continued dDAVP) and water-loaded (continued dDAVP plus a daily oral water load). A significant increase in urine volume in the water-loaded rats was observed by the second day of water loading, indicating onset of vasopressin escape. The onset of escape coincided temporally with a marked decrease in renal aquaporin-2 protein (measured by semiquantitative immunoblotting), which began at day 2 and fell to 17% of control levels by day 3. In contrast, there was no decrease in the renal expression of aquaporins 1, 3, or 4. The marked suppression of whole kidney aquaporin-2 protein was accompanied by a concomitant suppression of whole kidney aquaporin-2 mRNA levels. Immunocytochemical localization and differential centrifugation studies demonstrated that trafficking of aquaporin-2 to the plasma membrane remained intact during vasopressin escape. The results suggest that escape from vasopressin-induced antidiuresis is attributable, at least in part, to a vasopressin-independent decrease in aquaporin-2 water channel expression in the renal collecting duct.

Hyponatremia in rats induces downregulation of vasopressin synthesis.

Hyponatremia due to inappropriate secretion of vasopressin is a common disorder in human pathophysiology, but vasopressin synthesis during hypoosmolality has not been investigated. We used a new method to quantitate synthesis of vasopressin in rats after 3, 7, and 14 d of hyponatremia induced by administering dDAVP (a vasopressin agonist) and a liquid diet. Vasopressin synthesis was completely turned off by 7 d. Vasopressin mRNA levels in the hypothalamus paralleled the reduction in synthesis and were reduced to levels of only 10-15% of the content in control rats. When hyponatremia was corrected by withdrawal of dDAVP, vasopressin mRNA slowly returned to normal over 7 d. The observation that vasopressin synthesis can be so completely turned off leads to several conclusions: under normal physiological conditions the neurohypophysis is chronically upregulated; there must be an osmotic threshold for initiation of vasopressin synthesis (and release); the large store of hormone in the posterior pituitary is essential for vasopressin to be available during times of decreased synthesis; and, finally, some nonosmolar stimulus for synthesis must be present during clinical disorders when vasopressin is secreted (and synthesized) despite hypoosmolality.

Responses of magnocellular neurons to osmotic stimulation involves coactivation of excitatory and inhibitory input: an experimental and theoretical analysis.

How does a neuron, challenged by an increase in synaptic input, display a response that is independent of the initial level of activity? Here we show that both oxytocin and vasopressin cells in the supraoptic nucleus of normal rats respond to intravenous infusions of hypertonic saline with gradual, linear increases in discharge rate. In hyponatremic rats, oxytocin and vasopressin cells also responded linearly to intravenous infusions of hypertonic saline but with much lower slopes. The linearity of response was surprising, given both the expected nonlinearity of neuronal behavior and the nonlinearity of the oxytocin secretory response to such infusions. We show that a simple computational model can reproduce these responses well, but only if it is assumed that hypertonic infusions coactivate excitatory and inhibitory synaptic inputs. This hypothesis was tested first by applying the GABA(A) antagonist bicuculline to the dendritic zone of the supraoptic nucleus by microdialysis. During local blockade of GABA inputs, the response of oxytocin cells to hypertonic infusion was greatly enhanced. We then went on to directly measure GABA release in the supraoptic nucleus during hypertonic infusion, confirming the predicted rise. Together, the results suggest that hypertonic infusions lead to coactivation of excitatory and inhibitory inputs and that this coactivation may confer appropriate characteristics on the output behavior of oxytocin cells. The nonlinearity of oxytocin secretion that accompanies the linear increase in oxytocin cell firing rate reflects frequency-facilitation of stimulus-secretion coupling at the neurohypophysis.

MDL 17,043 therapy in severe congestive heart failure: characterization of the early and late hemodynamic, pharmacokinetic, hormonal and clinical response.

MDL 17,043, an agent with both inotropic and vasodilator properties, was evaluated in the treatment of chronic severe heart failure. The early and late hemodynamic, hormonal, pharmacokinetic and clinical responses to oral MDL 17,043 were studied in 20 patients. MDL 17,043 acutely increased cardiac output from 3.6 +/- 0.9 to 4.6 +/- 1.0 liters/min (+28%, p less than 0.001) and decreased mean pulmonary artery wedge pressure from 24 +/- 8 to 13 +/- 8 mm Hg (-46%, p less than 0.001), mean right atrial pressure from 10 +/- 5 to 4 +/- 4 mm Hg (-60%, p less than 0.001) and mean arterial pressure from 78 +/- 9 to 70 +/- 11 mm Hg (-10%, p less than 0.001). Hemodynamic improvement was sustained for 8 hours. Plasma renin activity tended to increase (0.10 less than p greater than 0.05), plasma norepinephrine tended to decrease (0.10 less than p greater than 0.05) and arginine vasopressin did not show any directional change. Elimination half-life for MDL 17,043 was approximately 20 hours. Hemodynamic responsiveness was maintained in six patients undergoing restudy at 4 weeks. Initial subjective improvement in the 20 patients occurred in 90%, was present at 4 weeks in 50% and continued longer than 3 months in 25%. Side effects occurred in 75% and required cessation of treatment in 10%. Thirteen (93%) of 14 patients on long-term therapy died (median time after start of MDL 17,043 therapy 39 days). Deaths were sudden in 69%. It is concluded that oral MDL 17,043 produces early and late hemodynamic improvement in patients with severe heart failure.(ABSTRACT TRUNCATED AT 250 WORDS)

Hyponatremia: endocrinologic causes and consequences of therapy.

Hyponatremia is a relatively common medical disorder frequently of endocrinologic origin. Although the neurologic signs and symptoms associated with hyponatremia are well known, recent clinical and experimental studies have shown that rapid correction of hyponatremia can in some cases cause a brain-demyelinating disease that can lead to consequences just as severe as those of hyponatremia itself. Understanding the physiologic mechanisms by which the brain adapts to hypoosmolar conditions has led to a better appreciation of the pathogenesis of neurologic dysfunction during both hyponatremia and its therapy, and this in turn has allowed the establishment of rational guidelines for safe therapy of hyponatremic patients. In cases such as this, however, in which a spontaneous rapid water diuresis occurs following a change in hormonal therapy, the rate and magnitude of the increase in serum [Na+J may exceed these guidelines unless active intervention is undertaken to interrupt the induced diuresis.

Vasopressin V2 receptor antagonists.

Hyponatremia due to the syndrome of inappropriate antidiuretic hormone secretion (SIADH) and disorders of water retention such as congestive heart failure and cirrhosis is a common problem encountered in the care of the medical patient. Thus far, available treatment modalities for disorders of excess arginine vasopressin (AVP) secretion or action have been suboptimal. The development of nonpeptide AVP V2 receptor antagonists represents a promising treatment option to directly antagonize the effects of elevated plasma AVP concentrations by increasing the water permeability of renal collecting tubules, thereby promoting excretion of retained water and normalizing hypoosmolar hyponatremia. In this review, SIADH and other water retaining disorders are briefly discussed, after which the published preclinical and clinical studies in the development of several nonpeptide AVP V2 receptor antagonists are summarized. The likely therapeutic indications and potential complications of these compounds, as well as their vascular effects, are also described.

Hyponatremia-induced inhibition of magnocellular neurons causes stressor-selective impairment of stimulated adrenocorticotropin secretion in rats.

Chronically hyponatremic rats were subjected to various stressors in order to evaluate the possible contribution of magnocellular neurons to the regulation of ACTH secretion, since such rats have markedly inhibited secretion and synthesis of magnocellular arginine vasopressin (AVP) and oxytocin (OT). Stress caused by a novel environment or by insulin-induced hypoglycemia resulted in moderate increases in plasma ACTH, which were of similar magnitude in both hyponatremic and normonatremic rats, and these stressors caused no increase in plasma AVP and OT levels in either group of rats. However, when exposed to ether, hyponatremic rats exhibited a significantly blunted ACTH response compared to normonatremic controls (331 +/- 49 vs. 740 +/- 124 pg/ml; P less than 0.01, respectively), and plasma AVP levels were markedly increased in the normonatremic, but not in the hyponatremic, rats. Intravenous infusion of 2 M NaCl also caused an ACTH release in hyponatremic rats that was significantly smaller than that in their normonatremic counterparts (228 +/- 52 vs. 479 +/- 85 pg/ml; P less than 0.05, respectively), and in this case both plasma AVP and OT levels were markedly increased in the normonatremic, but not in the hyponatremic, rats. However, hyponatremic rats exhibited greatly increased plasma ACTH levels 2 and 96 h after adrenalectomy (ADX), which were statistically equivalent to the increases in ACTH levels in normonatremic rats after ADX. Seven days after ADX parvocellular neurons of the paraventricular nucleus showed strongly increased CRF-41 and AVP-neurophysin, but not OT-neurophysin, immunoreactivities in both normonatremic and hyponatremic rats. These results show that parvocellular CRF-41/AVP-producing neurons in the paraventricular nucleus are not inhibited by chronic hyponatremia, in contrast to magnocellular neurons, and suggest that ACTH secretion induced by ether or hypertonic saline, but not by novel environment or insulin-induced hypoglycemia, is partially mediated by magnocellular AVP and/or OT.

Oxytocin produces natriuresis in rats at physiological plasma concentrations.

Oxytocin (OT) is known to stimulate natriuresis in rats when administered in large doses that produce high plasma levels. We examined the effects of physiological plasma OT levels on renal sodium excretion by infusing graded doses of OT sc in conscious adult male rats maintained on a sodium-deficient diet. Our results demonstrate that OT causes a dose-related increase in urinary sodium excretion during the initial day of infusion. The lowest plasma OT levels associated with increases in urinary sodium excretion (5-6 pmol/liter) were well within the range of physiological OT secretion in rats. However, this natriuretic effect was not sustained during subsequent days of maintenance on a sodium-deficient diet, suggesting that the OT-induced natriuresis was limited in part by receptor desensitization and/or a decreased exchangeable sodium pool in combination with secretion of opposing antinatriuretic factors such as aldosterone. Pretreatment with an OT receptor antagonist completely blocked the natriuresis produced by a 20 pmol/h infusion of OT, but urinary sodium excretion was not affected by a vasopressin V1 antagonist and was blocked only partially by a combined vasopressin V1 and V2 antagonist. Together with previous studies in rats demonstrating an inverse relation between pituitary OT secretion and sodium appetite, these results support the hypothesis that peripherally and centrally secreted OT act in concert in rats to produce a negative sodium balance by stimulating sodium excretion while inhibiting sodium ingestion.

Estradiol attenuates angiotensin-induced aldosterone secretion in ovariectomized rats.

Estrogen replacement therapy significantly reduces the risk of cardiovascular disease in postmenopausal women. Previous studies indicate that estradiol (E2) decreases angiotensin II (AT) receptor density in the adrenal and pituitary in NaCl-loaded rats. We used an in vivo model that eliminates the potentially confounding influence of ACTH to determine whether the E2-induced decrease in adrenal AT receptor expression affects aldosterone responses to angiotensin II (Ang II). Female rats were ovariectomized, treated with oil (OVX) or E2 (OVX+E2; 10 microg, s.c.) for 14 days, and fed a NaCl-deficient diet for the last 7 days to maximize adrenal AT receptor expression and responsiveness. On days 12-14 rats were treated with dexamethasone (DEX; 25 microg, i.p., every 12 h) to suppress plasma ACTH. On day 14 aldosterone secretion was measured after a 30-min infusion of Ang II (330 ng/min). Ang II infusion increased the peak plasma aldosterone levels to a lesser degree in the OVX+E2 than in the OVX rats (OVX, 1870 +/- 290 pg/ml; OVX+E2, 1010 +/- 86 pg/ml; P < 0.05). Ang II-induced ACTH and aldosterone secretion was also studied in rats that were not treated with DEX. In the absence of DEX, the peak plasma aldosterone response was also significantly decreased (OVX, 5360 +/- 1200 pg/ml; OVX+E2, 2960 +/- 570 pg/ml; P < 0.05). However, E2 also reduced the plasma ACTH response to Ang II (P < 0.05; OVX, 220 +/- 29 pg/ml; OVX+E2, 160 +/- 20 pg/ml), suggesting that reduced pituitary ACTH responsiveness to Ang II contributes to the effect of E2 on Ang II-induced aldosterone secretion. Adrenal AT1 binding studies confirmed that E2 significantly reduces adrenal AT1 receptor expression in both the presence and absence of DEX in NaCl-deprived rats. These results indicate that E2-induced decreases in pituitary and adrenal AT1 receptor expression are associated with attenuated pituitary ACTH and adrenal aldosterone responses to Ang II and suggest that estrogen replacement therapy may modulate Ang II-stimulated aldosterone secretion as part of its well known cardioprotective actions.

Characterization of oxytocin receptor expression and distribution in the pregnant sheep uterus.

At the end of pregnancy, the myometrium becomes extremely sensitive to oxytocin (OT) as result of a dramatic increase in the number of OT receptors (OTR), indicating an important role for OTR in the process of labor. There are no studies in sheep in which the physical properties and histological distribution of OTR are evaluated in relation to parturition. Also, no studies have been performed in any species to simultaneously examine the distribution of OTR at the messenger RNA (mRNA) as well as the protein levels in the same tissues and correlate those changes with the patterns of myometrial activity that occur at labor. In the present studies, we have used a polyclonal anti-OTR antibody and Western blot analysis to determine the apparent molecular mass of ovine OTR in late pregnant sheep myometrium and endometrium. We also examined the distribution of OTR mRNA and protein expression in the intact myometrium and endometrium and in individual cultured cells using in situ hybridization and immunocytochemistry. The expression of OTR and its mRNA has been correlated with the patterns of activity observed in the pregnant sheep myometrium. Western blot analysis of myometrial and endometrial extracts revealed a major form of OTR with an approximate molecular mass of 66 kDa. Both immunocytochemistry and in situ hybridization localized OTR and its mRNA in myometrial cells and glandular cells of the endometrium. Increased OTR and its mRNA expression in the myometrium and endometrium were correlated with the occurrence of myometrial contractions. OTR was also demonstrated by immunocytochemistry in the smooth muscle of myometrial blood vessels. Localization of OTR and its mRNA in pregnant sheep myometrial cells is consistent with the hypothesis that OTR plays an important role in regulating myometrial contractility. Positive staining of OTR in endometrial glandular cells supports the view that OT is involved in PG production by the endometrium in late pregnancy. Increased expression of OTR and its mRNA in the myometrium during labor further indicates that changes in tissue OTR play a significant role in the mechanism of parturition. Increased expression of OTR and its mRNA in endometrium may relate to the role of OT in regulating PG production by the endometrium during labor.

Cholecystokinin stimulates gonadotropin-releasing hormone release in the monkey (Macaca mulatta).

There is evidence to suggest that the arginine vasopressin release observed in association with emesis after i.v. injection of cholecystokinin (CCK) and N-methyl-D-aspartate (NMDA) is mediated by stimulation of emetic centers in the brainstem. That the GnRH-releasing action of NMDA is also sometimes accompanied by emesis led to the suggestion that stimulation of this parvocellular system by the glutamate agonist may also be mediated in part via activation of brainstem pathways. If this is the case, then other nauseogenic agents, such as CCK, should similarly elicit GnRH release. The foregoing prediction was tested in the castrated juvenile male monkey, an experimental model characterized by the absence of spontaneous GnRH release. GnRH discharges were monitored indirectly by measuring changes in circulating LH concentrations after the responsivity of the gonadotroph had been heightened by a chronic intermittent i.v. infusion of the decapeptide. An i.v. bolus of CCK at 10 and 30 micrograms/kg BW led to a distinct discharge of GnRH accompanied by emesis or other behaviors suggestive of nausea. Lower doses of CCK (1 and 3 micrograms/kg BW) failed to significantly perturb GnRH release or cause emesis, although NMDA (5 mg/kg BW; racemic form) injected i.v. 3 h after the CCK challenge led to a robust rise in GnRH. In a parallel study, three repetitive i.v. CCK injections at 2h intervals maintained intermittent GnRH release. Pretreatment with a long-acting GnRH receptor antagonist ([AcD2Nal1,4ClPhe2,DTrp3,DArg6,DAla10]GnRH -HOAc) abolished the LH response to CCK, confirming that the action of this peptide was mediated by GnRH release. Although a direct hypothalamic site of action for these agents remains the most likely possibility, since both NMDA and CCK receptors are present in the infundibular region, the present data are consistent with the notion that CCK and, by inference, NMDA may activate GnRH release in part via the stimulation of brainstem emetic centers. Plasma GH, PRL, and cortisol concentrations were also monitored during the course of some of these experiments, and the release of these hormones was observed after the administration of either the 10 or 30 micrograms/kg BW dose of CCK.

Vasopressin V2 receptor binding is down-regulated during renal escape from vasopressin-induced antidiuresis.

This study evaluated whether renal escape from vasopressin-induced antidiuresis is associated with alterations of vasopressin V2 receptor binding in the kidney inner medulla. A radioligand binding assay was developed using a novel iodinated vasopressin V2 receptor antagonist to analyze vasopressin V2 receptor binding in kidney inner medullary tissue from three groups of rats: normal rats maintained on ad libitum water intake, rats treated with 1-deamino-[8-D-arginine]vasopressin (DDAVP), and rats treated with DDAVP that were also water loaded to induce renal escape from antidiuresis. Analysis of the binding data showed that DDAVP treatment reduced vasopressin V2 receptor binding to 72% of normal levels. Water loading induced a marked further down-regulation of vasopressin V2 receptor binding. This receptor down-regulation began by day 2 of water loading, which correlated with the initiation of renal vasopressin escape; by day 3 of water loading, vasopressin V2 receptor expression fell to 43% of DDAVP-treated levels. No differences in vasopressin V2 receptor binding affinities were found among the three groups. This study demonstrates that vasopressin V2 receptor binding capacity is down-regulated during renal escape from vasopressin-induced antidiuresis and suggests that both vasopressin-dependent mechanisms as well as vasopressin-independent mechanisms associated with water loading are involved in this receptor down-regulation.

Estrogen regulates angiotensin AT1 receptor expression via cytosolic proteins that bind to the 5' leader sequence of the receptor mRNA.

Two of the most highly recognized factors implicated in the pathogenesis of hypertension, atherosclerosis, congestive heart failure and associated cardiovascular disease are the renin angiotensin system (RAS) and estrogen. A major effect of estrogen results from its influence on the RAS. Beta-estradiol (E2) replacement in ovariectomized (OVX) rats significantly decreased type 1 angiotensin (AT1) receptor expression in the pituitary and adrenal, whereas it significantly increased receptor expression in the uterus when compared to OVX controls. Additional evidence demonstrated an important influence of estrogen on a recently discovered post-transcriptional mechanism for regulating expression of the AT1 receptor. This mechanism consists of cytosolic RNA binding proteins (BPs) that recognize the 5' leader sequence (5'LS) of the receptor mRNA. The activities of these 5'LS BPs were modulated by estrogen in an inverse manner to AT1 receptor regulation. Moreover, in vitro translation assays in wheat germ lysates suggested that the 5'LS BPs inhibited AT1 receptor translation. Our data therefore indicate that hormonal regulation of AT1 receptors involves modulation of 5'LS BPs by estrogen. These findings may in part account for the observed protective effects of estrogen on cardiovascular disease.

Brain oxytocin receptor antagonism blunts the effects of anorexigenic treatments in rats: evidence for central oxytocin inhibition of food intake.

The inhibition of food intake in rats that results from various anorexigenic treatments is frequently associated with pituitary secretion of oxytocin (OT), but is not caused by circulating OT. We, therefore, evaluated the potential role of brain OT in mediating anorexia induced in rats by systemic administration of cholecystokinin (CCK), hypertonic saline (HS), or lithium chloride (LiCl), treatments that are known to stimulate pituitary OT secretion as well as to inhibit food intake. Food intake was analyzed in 22-h food-deprived rats pretreated with icv injections of either artificial cerebrospinal fluid (aCSF) or 9 nmol of an OT receptor antagonist, [d(CH2)5, Tyr(OMe)2,Orn8]vasotocin (OVT), which was the dose found to be most effective to antagonize the anorexia induced by CCK and HS. Pretreatment with the OT receptor antagonist icv significantly blunted the anorexigenic effect of each agent. After CCK (10 micrograms/kg, ip), food intake increased from 28 +/- 5% of basal intake after a CSF icv to 48 +/- 8% after OVT icv (P less than 0.01); after HS (2 ml 2 M NaCl, ip), food intake increased from 9 +/- 4% of basal intake after aCSF icv to 43 +/- 7% after OVT icv (P less than 0.01); and after LiCl (1.125 mmol/kg, ip), food intake increased from 55 +/- 4% of basal intake after a CSF icv to 80 +/- 9% after OVT icv (P less than 0.01). These data support the hypothesis that pituitary secretion of OT after anorexigenic treatments in rats is associated with coactivation of centrally projecting brain OT pathways, some of which are causally related to the induced inhibition of food intake.

Rat oxytocin receptor in brain, pituitary, mammary gland, and uterus: partial sequence and immunocytochemical localization.

Partial complementary DNAs of an oxytocin (OT) receptor were cloned from rat brain and uterus. The complementary DNAs encoded for the same amino acid sequence, which showed a high degree of homology with the human and porcine uterine OT receptors, except for a region in the third intracellular loop. Antibodies were raised against nonoverlapping sequences of the third intracellular loop of this rat OT receptor. Using these antisera, OT receptor expression was demonstrated in the brain, pituitary, mammary gland, and uterus by immunocytochemistry. In the brain, several areas including the ventromedial hypothalamus, the bed nucleus of the stria terminalis, the ventral pallidum, the paraventricular nucleus, and the dorsal part of the supraoptic nucleus, demonstrated OT-receptor immunoreactivity. However, no immunoreactivity was detected in two areas of the brain known to contain dense OT-binding sites by receptor autoradiography studies: the ventral hippocampus and the central nucleus of the amygdala. In the pituitary, both the anterior and posterior lobes were positive for OT receptor immunoreactivity, whereas the intermediate lobe was negative. These results demonstrate that the same receptor type is expressed in both peripheral OT target tissues and the brain, and also suggest the possibility that a different OT receptor subtype may be present in some areas of the brain.

Characterization of neurophysin-vasopressin prohormones in human posterior pituitary tissue.

To better characterize putative neurophysin-vasopressin prohormones in human posterior pituitary tissue, we extracted human posterior pituitary glands in 0.1 M HCl and isolated the higher molecular weight neurophysin-immunoreactive proteins. Sephadex G-75 gel filtration in 0.1 M formic acid with 6 M urea showed four distinct peaks of neurophysin immunoreactivity. Analysis of isolated lyophilized fractions of these peaks by sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed neurophysin-immunoreactive proteins at molecular weights of 10,000 daltons (79-87% of the total neurophysins), 19,000-20,000 daltons (10-16%), 26,000-30,000 daltons (1-2%), and a broad range of 30,000- to 100,000-dalton immunoreactivity from the void volume (V0) peak (2-3%). The 19,000- to 20,000-dalton and 26,000- to 30,000-dalton proteins were stable after both heating and treatment with reducing agents, but could be converted by chymotrypsin proteolysis to 10,000-dalton neurophysins and 3,000- to 5,000-dalton AVP-immunoreactive proteins. In contrast, the neurophysin immunoreactivity in the V0 peak was broken down to lower molecular weight neurophysin- and AVP-immunoreactive proteins by heating alone. Extraction of human posterior pituitaries in the presence of either [125I]human AVP-neurophysin or [35S] cysteine-labeled monkey neurophysin showed that no labeled neurophysin eluted in the areas of the 19,000- to 20,000- or 26,000- to 30,000-dalton proteins, but a significant fraction of the [35S]monkey neurophysin eluted in the V0. These data suggest that the 19,000- to 20,000- and 26,000- to 30,000-dalton human neurophysins represent stable proteins which are probably common precursor molecules for neurophysin and AVP, but the greater than 30,000-dalton neurophysins found in the V0 appear to be aggregates of neurophysins, neurophysin precursors, AVP, oxytocin, and probably other proteins and lipids as well, rather than very high molecular weight precursor proteins.