Effects of arginine vasotocin on levels of plasma gonadotropins and prolactin in ovariectomized conscious rats

Arginine vasotocin was injected into the third ventricle or intravenously in conscious, ovariectomized rats and its effect on gonadotropin and prolactin release evaluated. The peptide lowered plasma levels of both LH and prolactin in doses of 40 or 100 ng given intraventricularly. The higher dose was slightly more effective than the lower dose. Intravenous injection of a 1-microgram dose of vasotocin failed to alter plasma LH in the ovariectomized animals; however, a 5-micrograms dose induced a slight depression apparent at only 60 min following injection. Intravenous injection of 1 microgram produced a significant lowering of plasma prolactin, whereas a dramatic lowering followed the injection of the higher dose. Plasma FSH was unaffected in these experiments. Incubation of dispersed anterior pituitary cells from ovariectomized rats with various doses of vasotocin revealed no effect of the peptide on the release of FSH, LH, or prolactin. It also did not alter the response to LHRH, but it partially blocked the action of dopamine to inhibit prolactin release. The data indicate that quite low doses of arginine vasotocin act within the brain to inhibit LH and prolactin secretion in ovariectomized, conscious animals.     

Effects of changes in plasma hepatic-portal and systemic osmolality on plasma concentrations of arginine vasopressin in conscious newborn calves

Systemic plasma concentrations of arginine vasopressin (AVP) were studied in three groups of 10-15 day-old conscious newborn calves. Animals in the first group (control group) and in the second group (systemic-hypertonic-injected group) received respectively isotonic and hypertonic (8 mmol NaCl/kg body weight) saline injection into the right jugular vein. Animals in the third group were fitted with chronic mesenteric and hepatic-portal catheters and received a 1 h-hypertonic saline infusion (2 mmol NaCl/kg body weight) into the main mesenteric vein. In animals in the second group there were parallel increases in systemic plasma concentration of Na+ (from 148.0 +/- 2.6 to 177 +/- 8 mmol/l; P less than 0.01), osmolality (from 289 +/- 2 to 319 +/- 4 mOsmol/kg H2O; P less than 0.01) and systemic plasma concentrations of AVP (from 4.2 +/- 0.4 to 11.1 +/- 0.6 pmol/l; P less than 0.01) 10 min after the injection. There were no significant changes in control animals. Hypertonic saline infusion into the main mesenteric vein in the third group induced an increase in concentration of Na+ (from 147.3 +/- 2.0 to 165.0 +/- 5.0 mmol/l; P less than 0.01) and osmolality (from 288 +/- 5 to 315 +/- 10 mOsmol/kg H2O; P less than 0.01) in hepatic-portal vein plasma but did not alter systemic plasma osmolality or concentrations of Na+ and AVP. This study demonstrates that the relationship between plasma concentrations of AVP and systemic osmolality is operative in the newborn calf but does not support the hypothesis that hepatic portal osmo-receptors sensitive to hyperosmolality influence AVP release.     

Effect of atrial natriuretic factor on plasma vasopressin in conscious rats

An intravenous (IV) bolus injection (10 μg) of synthetic rat atrial natriuretic factor [ANF (Arg 101-Tyr 126)] into normal conscious Sprague-Dawley rats produced a significant decrease of plasma arginine vasopressin (AVP) while 1-, 2-, and 5-μg doses exerted no such effect. Mean arterial blood pressure (MAP) was lowered about 15 mmHg by an IV 10 μg bolus injection of ANF. When plasma AVP rose significantly in rats exposed to such osmotic stimuli as 600 mM NaCl and 900 mM mannitol intraperitoneally (IP), subsequent IV injection of ANF (10 μg) markedly depressed this parameter. Lower doses of ANF were ineffective against 600 mM NaCl IP. The significant elevation of plasma AVP levels by hypertonic sucrose 900 mM IP was not modified by ANF (10 μg). Blood pressure remained unchanged after IP administration of various osmotic stimuli, except mannitol, and in all these experiments an IV bolus of ANF exerted a lowering effect on MAP. Seventy-two hr water deprivation (mixed osmotic and volume stimulus) resulted in elevated plasma AVP levels which were unaffected by an IV bolus injection of ANF at doses of 0.06–10 μg. Immunoreactive ANF (IR-ANF) rose in plasma to 39.3±13 ng/ml 1 min after an IV bolus injection of 10 μg ANF, dropping to 1.01±0.2 ng/ml after 5 min and to 0.32±0.01 ng/ml after 10 min (when ANF and AVP interactions were studied), but still remained approximately six times higher than in control rats. These results suggest that, in the conscious rat, only pharmacological levels of ANF observed after an IV bolus infusion may influence both resting and osmotically-stimulated AVP levels.     

Radioimmunoassay of arginine vasopressin in human plasma.

Antibodies for the radioimmunoassay of arginine vasopressin (AVP) described here were produced in rabbits using synthetic AVP coupled to rabbit gamma-globulin with carbodiimide. In three out of six rabbits, significant antibody titres were obtained. Using the best antisera produced, 40% of labeled AVP was bound at a final dilution of 1:50.000. After iodination of synthetic AVP with 125I using the chloramin-T method, a gel filtration on Sephadex G-25 was performed to purify the iodinated AVP. For separation of antibody bound and free hormone, a second antibody precipitation was used. There was no crossreactivity with oxytocin. AVP was extracted from plasma after ammoniumsulfate precipitation of the proteins by adsorption to Florisil. The recovery of AVP added to plasma in amounts between 5-25 pg/ml was 60 +/- 15% (n equals 6). The minimum amount of AVP detectable was 1 pg per ml plasma. The plasma level in normal adults under standard conditions was 3.4 +/- 2.2 pg/ml. This is in agreement with data recently published by other researchers. The applicability and reproducibility was further tested in measurements of samples taken hourly during the entire day under water diuresis and after hormonal stimulation of AVP.     

Effects of treadmill running and swimming on plasma and brain vasopressin levels in rats

The influence of treadmill or swimming exercise on resting values of plasma and brain arginine vasopressin (AVP), and plasma sodium, potassium, osmolality and proteins was studied after 5 weeks of training using female Wistar rats. The duration of daily training sessions was progressively increased to reach 6 h/day for swim training (S) and 3 h/day for treadmill running (T). Compared to their untrained controls, treadmill and swim training were respectively associated with: a significant lower body weight; a decreased plasma AVP (36.4% for T and 47.4% for S) and hypothalamic AVP (20% for T and 16% for S); a higher hypophyseal AVP (145% for T and 36.3 for S); a decreased plasma osmolality (6.7% for T and 6.1% for S), sodium (1.2% for both) and potassium (15% for T and 22.4% for S); and no change in protein concentration. For T, rectal temperature increased (38.5 +/- 0.20 to 39.7 +/- 0.5) and for S rectal temperature decreased from 38.6 +/- 0.12 to 37.74 +/- 0.10). The differences observed in AVP contents of the pineal and Harderian glands (enhanced only in the treadmill groups) could be explained by the supposed role of these glands in thermoregulation. Two conclusions could be drawn from this study: there are no parallel changes in the hypothalamo-hypophyseal system (where AVP plays its endocrine role) and the brain (where AVP is a neurotransmitter); plasma changes could be explained by an extracellular fluid expansion with Na and K loss leading to a decrease in AVP secretion.     

Regulation of plasma vasopressin and renin activity in conscious hindlimb-unloaded rats

Cardiovascular deconditioning occurs in astronauts after spaceflight or in individuals subjected to bed rest. It is characterized by an increased incidence of orthostatic intolerance. The mechanisms responsible for orthostatic intolerance are likely multifactorial and may include hypovolemia, autonomic dysfunction, and vascular and cardiac alterations. The arterial baroreflex is an important compensatory mechanism in the response to an orthostatic stress. In a previous study, we demonstrated that arterial baroreflex mediated sympathoexcitation was blunted in hindlimb-unloaded (HU) rats, a model of cardiovascular deconditioning. The arterial baroreflex also contributes to the regulation of vasoactive hormones including vasopressin and angiotensin II. In the present study, we tested the hypothesis that the neurohumoral response to hypotension is also attenuated in rats after 14 days of hindlimb unloading. To test this hypothesis, the vasodilator diazoxide (15 or 25 mg/kg) or saline (0.9%) was administered to produce hypotension or control conditions, respectively, in conscious HU and control rats. Plasma samples were collected and assayed for vasopressin and plasma renin activity (PRA). Diazoxide (25 mg/kg) produced significant increases in vasopressin and PRA compared with saline controls. HU rats exhibited significantly higher levels of vasopressin at rest and the increase in vasopressin levels during hypotension was enhanced by hindlimb unloading. Neither resting nor hypotension-induced PRA was altered by hindlimb unloading. These data suggest that although baroreflex-mediated sympathoexcitation is blunted by hindlimb unloading, hypotension-induced vasopressin release is enhanced and hypotension-induced PRA is unaffected. Increased circulating vasopressin may serve to compensate for blunted baroreflex regulation of sympathetic nervous activity produced by hindlimb unloading or may actually contribute to it.     

Electroacupuncture at ST-36 accelerates colonic motility and transit in freely moving conscious rats

Acupuncture is useful for functional bowel diseases, such as constipation and diarrhea. However, the mechanisms of beneficial effects of acupuncture on colonic function have scarcely ever been investigated. We tested the hypothesis that electroacupuncture (EA) at ST-36 stimulates colonic motility and transit via a parasympathetic pathway in conscious rats. Hook-shaped needles were inserted at bilateral ST-36 (lower limb) or BL-21 (back) and electrically stimulated at 10 Hz for 20 min. We also studied c-Fos expression in response to EA at ST-36 in Barrington's nucleus of the pons. EA at ST-36, but not BL-21, significantly increased the amplitude of motility at the distal colon. The calculated motility index of the distal colon increased to 132 +/- 9.9% of basal levels (n = 14, P < 0.05). In contrast, EA at ST-36 had no stimulatory effects in the proximal colon. EA at ST-36 significantly accelerated colonic transit [geometric center (GC) = 6.76 +/- 0.42, n = 9, P < 0.001] compared with EA at BL-21 (GC = 5.23 +/- 0.39, n = 7). The stimulatory effect of EA at ST-36 on colonic motility and transit was abolished by pretreatment with atropine. EA-induced acceleration of colonic transit was also abolished by extrinsic nerve denervation of the distal colon (GC = 4.69 +/- 0.33, n = 6). The number of c-Fos-immunopositive cells at Barrington's nucleus significantly increased in response to EA at ST-36 to 8.1 +/- 1.1 cells/section compared with that of controls (2.4 +/- 0.5 cells/section, n = 3, P < 0.01). It is concluded that EA at ST-36 stimulates distal colonic motility and accelerates colonic transit via a sacral parasympathetic efferent pathway (pelvic nerve). Barrington's nucleus plays an important role in mediating EA-induced distal colonic motility in conscious rats.     

A self-calibrating telemetry system for measurement of ventricular pressure-volume relations in conscious, freely moving rats

Using Bluetooth wireless technology, we developed an implantable telemetry system for measurement of the left ventricular pressure-volume relation in conscious, freely moving rats. The telemetry system consisted of a pressure-conductance catheter (1.8-Fr) connected to a small (14-g) fully implantable signal transmitter. To make the system fully telemetric, calibrations such as blood resistivity and parallel conductance were also conducted telemetrically. To estimate blood resistivity, we used four electrodes arranged 0.2 mm apart on the pressure-conductance catheter. To estimate parallel conductance, we used a dual-frequency method. We examined the accuracy of calibrations, stroke volume (SV) measurements, and the reproducibility of the telemetry. The blood resistivity estimated telemetrically agreed with that measured using an ex vivo cuvette method (y=1.09x - 11.9, r2= 0.88, n=10). Parallel conductance estimated by the dual-frequency (2 and 20 kHz) method correlated well with that measured by a conventional saline injection method (y=1.59x - 1.77, r2= 0.87, n=13). The telemetric SV closely correlated with the flowmetric SV during inferior vena cava occlusions (y=0.96x + 7.5, r2=0.96, n=4). In six conscious rats, differences between the repeated telemetries on different days (3 days apart on average) were reasonably small: 13% for end-diastolic volume, 20% for end-systolic volume, 28% for end-diastolic pressure, and 6% for end-systolic pressure. We conclude that the developed telemetry system enables us to estimate the pressure-volume relation with reasonable accuracy and reproducibility in conscious, untethered rats.     

Real-time cortical cerebral blood flow follow-up in conscious,freely moving rats by laser Doppler flowmetry

This article describes a laser Doppler flowmetry (LDF) system that enables repeated measurements and thereby long-term followup of cortical cerebral blood flow (CBF) in awake and freely moving rats. The system consists of a specially designed flow probe adapter, a flow probe connector, and a LDF flow probe, which may thereby rotate through its own axis. During the experiment, the flow adapter is permanently mounted onto the rat's skull bone. A thin layer of skull bone is left intact at the site for cortical CBF measurements. The probe connector and the flow probe may be repeatedly detached and remounted to the adapter, which allows for cortical cerebral blood flow recording from exactly the same anatomical location. The laser Doppler flowmetry system enables stable cortical CBF recordings in the conscious rat while it moves freely in a bowl cage.     

Metabolism and synthesis of arginine vasopressin in conscious newborn sheep

Arginine vasopressin (AVP) is an important regulator of cardiovascular homeostasis in the fetus, but its role after birth is unclear. Although infused AVP increases mean arterial pressure (MAP) during the 1st mo after birth, pressor responses are unchanged, suggesting that vascular responsiveness is also unchanged. Alternatively, this could reflect increases in AVP metabolic clearance rate (MCR(AVP)). However, newborn AVP metabolism and synthesis are poorly studied. Therefore, we examined the pressor responses to infused AVP and the pattern of circulating AVP, AVP production rate (PR(AVP)), and MCR(AVP) in conscious newborn sheep (n = 5) at 9-38 days after birth. Basal MAP rose and heart rate (HR) fell during the study period (P < or = 0.02), while circulating AVP was unchanged (P > 0.1), averaging 3.01 +/- 0.86 pg/ml. Infused AVP elicited steady-state responses at 10-40 min, increasing plasma AVP and MAP and decreasing HR (P < 0.001). Although pressor responses were unchanged between 9 and 38 days, the rise in MAP correlated with increases in plasma AVP (R = 0.47, P = 0.02, n = 24). MCR(AVP) was unchanged throughout the 1st mo (P > 0.2), averaging 205 +/- 17 ml.kg(-1).min(-1), and was associated with an elevated PR(AVP), 973 +/- 267 pg.kg(-1).min(-1), which also was unchanged (P > 0.1). After birth, MCR(AVP) and PR(AVP) are elevated, probably accounting for the stable plasma AVP levels. The former is also likely to account for the stable pressor responses to infused AVP during the 1st mo. The reason for the elevated PR(AVP) is unclear but may relate to increases in vascular volume associated with postnatal growth.     

Effect of intraventricular oxytocin and vasopressin on self-stimulation in rats.

Oxytocin (500 mu u) and vasopressin (50 mu u) were injected into the lateral ventricle and its effect on hypothalamic self-stimulation has been studied. Oxytocin increased, while vasopressin decreased the self-stimulation rate tested 10-20 min following application. The hypothalamic and mesencephalic serotonin content decreased slightly while plasma corticosterone content did not change 20 min after oxytocin and vasopressin administration compared to the injected control animals. The data suggest that vasopressin and oxytocin have an opposite effect on self-stimulation and this action is not mediated through the brain serotoninergic or pituitary-adrenocortical axis.     

Electric footshocks differentially affect plasma and spinal cord vasopressin and oxytocin levels

Rats exposed for three minutes to repeated electric footshocks showed an approximate 10-fold increase of basal plasma vasopressin (AVP) and oxytocin (OXT) levels. In contrast, spinal AVP and OXT contents measured in the same rats remained unchanged when compared to undisturbed controls. This observation suggests that spinal AVP and OXT do not play a major role in the short-term adaptation of the organism to stress.     

Effect of TRH on plasma arginine vasopressin

Effect of thyrotropic releasing hormone (TRH) on plasma arginine vasopressin (AVP) was studied in human subjects. All 7 normal controls and 2 hypothyrotropic hypothyroid subjects failed to show any rise of AVP on TRH administration. The 4 primary hypothyroid subjects had elevated basal AVP level and showed further elevation on TRH administration. Our data suggests that elevated TRH in primary hypothyroid subjects may act directly as a nonosmotic stimulus or modulate the osmoreceptor and hypothalamic neurohypophyseal system for AVP release.     

Radioimmunoassay of arginine vasopressin in the plasma and urine.

We introduced the radioimmunoassay (RIA) of arginine vasopressin (AVP) with standard AVP and antiserum to AVP (both Calibiochem). The sensitivity of the system was increased from the declared 4pg to 1 pg per tube by preparing AVP-125I of high specific activity (about 1,500 mCi/mg) and by modifying the reaction conditions. The sensitivity of the method was adequate for measuring AVP in urine and in concentrated plasma extracts, even under physiological conditions. Reliability of the results depended upon maintenance of approximately the same osmolarity in all the RIA samples. The mean plasma AVP level, uncorrected for AVP extraction losses, was 1.52 +/- 0.20 pg/ml for an ad libitum fluid intake; in fluid deprivation it rose in proportion to the osmolarity of the plasma to 5.83 +/- 0.42 pg/ml at 12 hours and to 19.09 +/- 4.51 pg/ml at 36 hours. Extraction recovery of added AVP was about 63%. The urinary AVP concentration varied according to the patients' state of hydratation from undetectable values at UOsm less than 200 mOsm/1 to a mean 16.5 +/- 7.9 pg/ml in the presence of an ad libitum fluid intake and to 29.1 +/- 7.5 pg/ml after 12 hours' and 117.2 +/- 13.7 pg/ml after 36 hours' deprivation of fluids.