Suppression by naloxone of water intake induced by deprevation and hypertonic saline in intact and hypophysectomized rats

Naloxone, an opiate antagonist, was administered to intact and hypophysectomized male rats following hypertonic saline pretreatment or 12 hr water deprivation. Water intake following hypertonic saline or water deprevation was reduced by 0.01 – 10 mg/kg of naloxone in a dose-related fashion in both intact and hypophysectomized rats. Water consumption induced by hypertonic saline administration appeared to be more susceptible to the suppressant effects of naloxone than did that evoked by water deprevation. These results demonstrate that naloxone reduces water intake in the rat following intracellular dehydration by hypertonic saline administration, as well as after general dehydration induced by water deprevation. Furthermore, the suppressant effects of naloxone on water intake do not appear to involve pituitary endorphins, although a possible involvement of antidiuretic hormone in these effects cannot be excluded.     

Brain enkephalin distribution is unaltered by hypophysectomy

Methionine (met) and leucine (leu)-enkephalin were measured by radioimmunoassay in 16 brain regions removed by microdissection in intact rats. In hypophysectomized rats the regional concentrations of the enkephalins was not altered. Brain enkephalins do not appear to be derived from pituitary endorphins.     

Pituitary opioid involvement in ECS-postictal electrogenesis and behavioral depression in rats

The role of pituitary opioids in electroconvulsive shock (ECS)-induced postictal electrogenesis and behavioral depression was investigated in sham-hypophysectomized and hypophysectomized rats. These animals were divided into two subgroups and injected SC with either saline or naloxone (3 mg/kg) 10 min prior to transauricular ECS. Sham-hypophysectomized rats given saline responded to a single ECS with a 65 +/- 18% (s.e.) increase in postictal electrogenesis and a behavioral depression lasting 3840 +/- 530 sec. Naloxone significantly antagonized both the postictal increase in EEG voltage output and behavioral depression. Hypophysectomy by itself was without effect on EEG patterns and only partially attenuated the ECS-induced electrogenesis and postictal depression (31.9 +/- 9% and 2360 +/- 511 sec, respectively). However, in hypophysectomized rats, naloxone did not further antagonize these effects of ECS. Thus, it appears that pituitary opioids may, at least in part, mediate postictal electrogenesis and behavioral depression. Alternatively, since hypophysectomy only partially attenuates these phenomena, central or nonpituitary opioid peptide systems may be involved. In view of the observed decrease in responsiveness to naloxone in hypophysectomized rats, nonopioid systems cannot be ruled out as contributors to the opioid-like effects of ECS in these animals.     

The role of the pituitary in the diurnal variation in tolerance to painful stimuli and brain enkephalin levels

Studies were carried out on hypophysectomized rats and mice in comparison to sham-operated controls in order to assess the role of the pituitary in the diurnal rhythm in sensitivity to pain, the hyperalgesic effect of naloxone and the effect of stress on brain levels of met-enkephalin. There were no significant differences in jump latencies between hypophysectomized and sham-operated control mice. The jump latencies in the p.m. were significantly greater than those in the a.m. for both the sham and the hypophysectomized mice. In both the sham and hypophysectomized mice and rats, naloxone significantly reduced the jump latencies in the p.m. The stress-induced increase in the p.m. of brain met-enkephalin, furthermore, persisted in the hypophysectomized rats. We conclude that the pituitary is not essential for the diurnal variation in responsivity to pain, the hyperalgesic activity of naloxone or the stress-induced increases in brain met-enkephalin.     

Neurally mediated gastric mucosal damage in hypophysectomized rats.

The role of the pituitary hormones in the development of neurally mediated gastric mucosal damage was examined in both normal and hypophysectomized urethane-anaesthetized male Sprague-Dawley rats. Gastric mucosal damage was elicited either by electrical stimulation of intact vagal nerves or by electrical stimulation in the paraventricular nucleus. Macroscopic damage was scored following the stimulation period and samples of the stomach were fixed for histological assessment. Damage scores were assigned based on a 0 (normal) to 3 (severe) scale. Control experiments in which the vagi were not stimulated did not result in any significant gastric damage in either normal (0.56) or sham surgery (0.14) animals, whereas hypophysectomized animals were observed to have significant damage (1.44, p < 0.05). Stimulation of the vagi in hypophysectomized animals resulted in damage that was not significantly different compared with the hypophysectomized control animals (1.25, p > 0.05). In normal animals, stimulation of vagal nerves resulted in mean damage scores of 2.00, values that were not significantly different from those observed in hypophysectomized animals (1.25, p > 0.05). Similarly, stimulation in the paraventricular nucleus of hypophysectomized animals resulted in gastric lesions (2.00) that were not significantly different from those observed in normal animals (1.91, p > 0.05). These data suggest that such neurally mediated gastric damage does not depend upon neurosecretory projections to the pituitary gland, but that the maintenance of an intact gastric mucosa under normal conditions requires the presence of pituitary hormones.     

Failure of hypophysectomy to alter brain content of opioid peptides (endorphins).

Opioid peptides (endorphins) were extracted from brains of hypophysectomized rats and sham-operated controls on the 34th post-operative day. There were no significant differences in endorphin content between groups. Evidently, the brain endorphins do not come from the pituitary, but are probably synthesized in the brain.     

Lysosomal enzymes in the rat harderian gland are altered by either bromocriptine treatment or hypophysectomy and hormone replacement therapy

Four groups of adult male hypophysectomized rats were injected subcutaneously twice daily between 0800-0900 hr and 1600-1700 hr with either saline diluent, 150 micrograms sheep prolactin and/or growth hormone (GH); intact rats received either saline or 150 micrograms bromocriptine twice daily. After 4 days of treatment, lysosomal enzyme assays revealed significant elevations in both acid phosphatase and alpha-mannosidase enzyme activities in the Harderian glands of saline-injected hypophysectomized rats compared to those in intact controls. beta-Glucuronidase levels were depressed and hexosaminidase activity unaffected by hypophysectomy treatment alone compared to intact controls. Lysosomal enzyme activities in hypophysectomized animals treated with prolactin were not different from the hypophysectomized control animals. However, treatment with GH alone or in combination with prolactin had a significant inhibitory effect on beta-glucuronidase, hexosaminidase, and alpha-mannosidase enzyme activities in the Harderian gland of hypophysectomized animals. Bromocriptine treatment in intact rats only elevated acid phosphatase activity. In summary, the patterns of responses did not reveal a role for prolactin in the control of Harderian gland lysosomal enzyme activities by the pituitary. However, some of the influence on this target system may be exerted by growth hormone.     

Endotoxin shock: prevented by naloxone in intact but not hypophysectomized rats

Previous studies established that naloxone reverses hypotension in endotoxin, hemorrhagic, and spinal shock. We studied endotoxin shock in hypophysectomized (Hx) rats, which have little circulating beta-endorphin. Hx or intact rats received surgically implanted jugular catheters for drug injection and aortic catheters for arterial blood pressure (MAP) recording. On the second day after implantation, rats were pretreated with either naloxone or saline. Two minutes later each rat received endotoxin. Following endotoxin, all rats showed a brief biphasic hypertensive-hypotensive response followed by stabilization of MAP near baseline. Within 20 min, all Hx rats, regardless of pretreatment, and the saline-treated intact rats, showed progressive hypotension (P less than 0.005). Only the naloxone-pretreated intact rats maintained a stable MAP. Plasma endorphin measured at 20 min was undetectable in Hx rats in contrast to intact rats (P less than 0.001); plasma corticosterone levels were likewise suppressed in the Hx rats (P less than 0.01). Thus (1) naloxone protected only the rats with an intact pituitary-adrenal-sympathetic system, and (2) pituitary endorphin is not required to generate endotoxin shock in hypophysectomized rats.     

Effects of growth hormone and hypophysectomy of pregnant rats on serum concentrations of pregnancy-associated murine protein-1

Continuous infusion of bovine GH to hypophysectomized non-pregnant rats increased serum concentrations of pregnancy-associated murine protein-1 (PAMP-1) to the levels of adult female rats and pregnant rats. Serum concentrations of PAMP-1 were followed from Day 16 of gestation until 3 days after parturition in hypophysectomized (on Day 14 of gestation) and intact pregnant rats. In the intact pregnant rat there was a decrease in PAMP-1 values from Day 16 until delivery. The serum concentrations of PAMP-1 in hypophysectomized pregnant rats were similar to those in intact pregnant rats before parturition, but PAMP-1 concentrations decreased markedly after parturition in the hypophysectomized rats. We suggest that the serum concentrations of PAMP-1 can be maintained without pituitary GH in late pregnancy, while serum values of PAMP-1 in non-pregnant rats is dependent upon a continuous secretion of pituitary GH.     

Diurnal variations in estradiol binding site concentrations in the hypothalamus, pituitary and uterus of prepuberal rats

Available cytosolic estradiol binding sites (EBS) were estimated at 6 h intervals over a 24 h period in individual extracts of uteri, pituitaries and hypothalami from intact, ovariectomized or hypophysectomized 28-day old rats, maintained from birth in a 12 h light-12 h dark photoperiod. Plasma estrogen levels were also recorded, in intact rats, they displayed a 20% rise during daytime. In the uterus and pituitary of ovariectomized rats EBS concentrations exceeded by one third those of intact controls. In intact animals, significant diurnal variations in EBS were observed in both these tissues, with increased daytime levels. These variations appeared to depend on the amount of estrogen in the plasma, as they were not observed in spayed rats. No diurnal variations in EBS were apparent in the hypothalamus of intact animals, but they did occur in ovariectomized rats, and even more significantly in hypophysectomized rats, although the latter had lower EBS concentrations than spayed animals. These variations were presumed to result from the interference of pituitary-controlled hormones other than estrogens in the regulation of EBS levels.     

Effect of prolactin and estradiol on rat striatal dopamine receptors

Chronic estrogen treatment has been found to increase the level of rat striatal dopamine receptors. Since it is well known that estrogen treatment increases circulating prolactin levels, we have investigated the possibility that the stimulatory effect of estrogens on dopamine receptors is exerted via prolactin. Ovariectomized female or intact male rats were implanted with three adenohypophyses under the kidney capsule or treated with 17 β-estradiol (10 μg, twice daily) for 2 weeks. In animals of both sexes, the pituitary-implanted and estradiol-treated rats showed higher levels of [³H]spiperone binding to striatal dopamine receptors. This effect of estradiol or pituitary implants on dopamine receptors was further investigated in ovariectomized rats. The pituitary-implanted and estradiol-treated rats had elevated plasma prolactin levels and an increased density of striatal dopamine receptors without alteration of their affinity. The role of the pituitary in the effect of estradiol was next investigated using hypophysectomized female rats treated with 17 β-estradiol (10 μg, twice daily), o-prolactin (500 μg, twice daily) or bearing three anterior pituitary implants. The implants as well as the treatment with estradiol or prolactin increased the level of striatal dopamine receptors in hypophysectomized rats while, as expected, the estradiol-treated animals did not have elevated plasma prolactin levels. The present data indicate that high prolactin levels lead, as observed with chronic estradiol treatment, to an increased density of striatal dopamine receptors. However, the effect of estradiol may not be explained exclusively by increased prolactin levels since a similar stimulatory effect is observed in hypophysectomized animals.     

The effect of hypophysectomy upon cholinesterase activity in the forelimbs of regenerating adult Notophthalmus news

In order to study endocrine influence upon cholinesterase activity during regeneration, adult newts were hypophysectomized either prior to limb transection or during regeneration. Homogenates of limb tissues were assayed for cholinesterase activity during each stage of regeneration.In animals with pituitaries intact, cholinesterase activity in regenerating limb tissues decreases soon after amputation, and then it rises to the level of activity in intact limbs of normal animals, during the period of differentiation. In hypophysectomized newts there seems to be no alteration of this basic pattern of activity, but removal of the pituitary does result in more elevated levels of enzymatic activity. In the intact forelimbs of control newts undergoing regeneration, cholinesterase activity greatly increases as the other transected limb begins to regenerate but it returns to normal as regeneration progresses. If these animals are hypophysectomized, no such increase is observed during the early stages of regeneration. Rather, there is an initial decrease in cholinesterase activity that is followed by an increase in such activity.These data are compatible with the hypothesis that the pituitary modulates cholinesterase activity in the limb tissues of adult newts.     

The concentration of atrial natriuretic peptide (ANP) is decreased in plasma but not in atria in hypophysectomised rats

To study the role of the pituitary gland in the release of Atrial Natriuretic Peptide (ANP) plasma and atrial concentrations were measured both in intact and in hypophysectomized rats. The plasma concentration of ANP (pg/ml) was significantly (p less than 0.01) decreased from 143 +/- 35 to 82 +/- 29 (mean +/- SD, n) while the tissue concentration (ng/wet tissue mg) remained unchanged, 192 +/- 46 and 194 +/- 39, respectively. The total atrial amount of ANP (ug) was, however, significantly (p less than 0.01) decreased from 29.7 +/- 7.8 to 17.0 +/- 3.3 after hypophysectomy. In intact animals, a volume load (1.1ml/100 body weight g 0.9% NaCl) resulted in 2-fold (p less than 0.001) increase in the plasma ANP levels whereas similar load had no effects on plasma ANP levels in hypophysectomized animals. In both groups, the right atrial pressure was increased from about 2 to about 6 mmHg. We conclude that in the absence of pituitary gland the right atrial pressure and the atrial ANP concentration do not change but plasma ANP levels and the response to volume stimulus are attenuated.     

Reactions of the adrenal cortex and thyroid gland of hypophysectomized rats to hypothermia

Male Wistar rats were hypophysectomized 7 days (group 1) and 4-7 weeks (group II) before exposure to hypothermia (4 degrees C for 1 1/2 h). The hypophysectomized rats from group I were devoid of both the posterior lobe and the adenohypophysis, while the rats from group II had the posterior hypophysis but not the adenohypophysis regenerated. A decreased arginine-vasopressin (AVP) blood level in group I (32%) and a very high level of AVP in group II (311%, P less than 0.05) was determined by RIA. The exposure to hypothermia did not influence the AVP plasma level. The thyroid hypofunction was revealed morphometrically in both hypophysectomized groups. Nevertheless, cooling stimulated the thyroid glands in rats of both experimental groups, like it was in the control. Thus, there is no evidence that thyroid gland reaction to hypothermia is affected by AVP. Cooling caused an increase of corticosteroid blood and adrenal cortex content in nonoperated control rats as well as in group II, but not in group I of experimental animals. Hence, it may be assumed that when the adenohypophysis is ablated, a high AVP blood level is necessary to realize the adrenal cortex response to hypothermia.     

The role of the pituitary in modifications of the uterine secretion of spayed, oestradiol-primed rats

Tests performed on spayed, adult female estradiol-primed Ivanovas rats, with ligated uteri and normal pituitary function have shown that treatment with sexual steroids, including progesterone and testosterone, modifies uterine secretion. One half of the animals were hypophysectomized. In estradiol primed hypophysectomized controls, growth was retarded about 28%, the weight of the empty uterus reduced, and the quantity of uterine secretion diminished in comparison with the values for the nonhypophysectomized controls. In test rats treated with estradiol, gain in body weight was virtually arrested in the nonhypophysectomized rats and a reduction in weight was observed in both groups treated with the highest dose of estradiol tested (300 mcg/kg daily). In rats treated with progesterone, no significant differences were found between the two groups. In treated groups, a dose-related reduction in the weight of the empty uterus was found. Treatment caused a marked reduction in the quantity of the uterine secretion, the effect appearing greater in nonhypophysectomized rats. Increasing doses of progesterone produced a rapid rise in the viscosity of the uterine fluid, as well as a decrease in the pH of the uterine lumen. In both hypophysectomized and nonhypophysectomized rats, testosterone induced a dose-related increase in body weight, statistically significant only in animals with intact pituitaries treated with 100 mg/kg daily. The weight of the empty uterus also increased. The quantity of uterine fluid was reduced by testosterone only when it was given in massive doses to nonhypophysectomized rats. Doses of 100-300 mg/kg daily were needed to produce the same response as a dose of about 10 mg/kg daily of progesterone. In response to large doses, viscosity of secretion rose slightly and the pH of uterine lumen and secretion decreased. It may be concluded that the progestative modifications induced by progesterone in the uterus of spayed, estradiol-primed rats, including particularly changes in uterine secretion, are the effects of a peripheral mechanism not involving the pituitary. Testosterone appears to be an exception as far as the quantity and viscosity of uterine secretion are concerned, since modifications in these parameters are only observed in the presence of a functional pituitary body.     

Nitrous oxide-induced hypothermia in the rat

Exposure of rats to high levels of nitrous oxide (N2O) in oxygen (O2) reduced body temperature in a concentration-related manner. The hypothermia was partly reversed by pretreatment with naloxone but not naltrexone. But in rats rendered tolerant to morphine by pellet implantation, exposure to 75% N2O/25% O2 evoked a marked hypothermia similar to that observed in morphine-naive animals. In another experiment, the hypothermic effect of chloral hydrate was also sensitive to antagonism by pretreatment with naloxone but not naltrexone. These observations lead us to suspect that N2O-induced hypothermia in rats is possibly not mediated by opiate receptors. The thermotropic activity of N2O may result from some non-opioid action of N2O. Its selective antagonism by naloxone (but not naltrexone) may be due to a unique non-opioid analeptic action of naloxone.     

Effects of pregnenolone-16 alpha-carbonitrile on drug metabolizing enzymes in hypophysectomized female rats

Treatment of intact and hypophysectomized female rats with pregnenolone-16 alpha-carbonitrile (PCN) resulted in a significant increase in hepatic aryl hydrocarbon hydroxylase (AHH) activity. However, the total cytochrome P-450 concentration, as measured by CO difference spectra, was increased to a greater extent in hypophysectomized rats than in intact rats. Total cytochrome P-450 was found to be 0.82 +/- 0.16 vs 2.43 +/- 0.31 nmoles/mg protein for control and PCN-treated hypophysectomized rats, respectively, and 0.68 +/- 0.23 vs 1.28 +/- 0.05 nmoles/mg protein for control and PCN-treated intact rats respectively. The concentration of metyrapone complex in microsomes from intact control and PCN-treated rats was found to be 0.4 +/- 0.11 vs 1.88 +/- 0.23 M respectively. Treatment of hypophysectomized rats with PCN resulted in an approximate 10-fold increase in the concentration of the metyrapone complex (0.42 +/- 0.15 M for control and 4.46 +/- 0.44 M for PCN-treated). Microsomal NADPH and NADPH cytochrome c reductase activities were also altered by PCN-treatment. Aminopyrine demethylase activity was stimulated approximately three-fold by PCN treatment in both intact and hypophysectomized rats. Benzphetamine demethylase activity was not significantly affected by PCN treatment. The results of these studies suggest that the absence of the pituitary gland can markedly influence PCN induction of cytochrome P-450 in the liver in female rats. PCN also differentially affects microsomal mixed-function oxidase activities associated with drug and xenobiotic metabolism.     

Divergence of growth hormone actions on hepatic drug metabolism in the presence and absence of the pituitary gland.

Experiments were carried out to compare the effects of growth hormone on hepatic drug oxidation in normal and hypophysectomized rats. Administration of growth hormone to normal male rats lowered hepatic microsomal cytochrome P-450 content and decreased the rates of ethylmorphine n-demethylation and aniline hydroxylation. These effects were fully manifested in orchiectomized or adrenalectomized males, excluding a dependence upon endogenous steroids. Growth hormone was without effect on hepatic drug metabolism or cytochrome P-450 content in normal female rats. In contrast to its actions in animals with intact pituitary glands, administration of growth hormone to hypophysectomized rats of either sex increased the rate of ethylmorphine metabolism. Furthermore, in both males and females, aniline hydroxylation and microsomal cytochrome P-450 content were unaffected by growth hormone in the absence of the pituitary gland. Prolactin administration did not affect hypophysectomized or in normal rats of either sex. The results indicate that the nature of growth hormone actions on hepatic drug oxidation is pituitary-dependent and probably intertwined with the effects of other hormones. Furthermore, the direct physiological effects of growth hormone on hepatic mixed function oxidases seem to depend upon the substrate employed.     

The luteinizing hormone-releasing hormone (LHRH) agonist, [D-Trp6, des-Gly-NH2(10)]-LHRH ethylamide, has no direct effect on spermatogenesis in the rat

Treatment of intact rats with luteinizing hormone-releasing hormone (LHRH) agonists has been shown to produce atrophy of a variable number of testicular seminiferous tubules. These findings raised the question of a possible direct versus indirect action of LHRH agonists on spermatogenesis. To answer this question, we treated hypophysectomized rats with the LHRH agonist [D-Trp6, des-Gly-NH2(10)]-LHRH ethylamide, dihydrotestosterone (DHT), or a combination of these two compounds for a period of 1 mo. Treatment of hypophysectomized animals with the LHRH agonist alone had no significant effect on the atrophy of seminiferous tubules found after hypophysectomy. DHT, however, maintained spermatogenesis at 80% of the level seen in intact animals. When DHT and the LHRH agonist were administered in combination, the stimulatory effects of DHT were observed with no significant interference caused by the LHRH agonist. This study shows that an LHRH agonist has no direct effect on the morphology of the seminiferous tubules in the absence of the pituitary gland and strongly suggests that the atrophy observed in the testis after LHRH agonist treatment in intact animals is mediated by the LHRH agonist-induced changes in luteinizing hormone secretion and/or direct action of the peptide on Leydig cells.     

Effects of thyroxine and cold exposure on hypothalamic TRH levels in rats with various pituitary-thyroid states.

The hypothalamic content and concentration of thyrotropin-releasing hormone (TRH) were determined by radioimmunoassay in normal, thyroidectomized, hypophysectomized and cold-exposed rats with or without thyroxine. In normal animals, the single administration of thyroxine (1,5 and 20 microgram/100 g B.W.) altered neither the content nor the concentration of TRH in the hypothalamus. However, seven days' administration of this hormone resulted in the dose-dependent increase in the hypothalamic TRH levels. In thyroidectomized rats the hypothalamic TRH levels were slightly reduced in spite of the marked increase of plasma TSH levels and decrease of pituitary TSH levels. In the animals given thyroxine (10 microgram/100 g B.W.) for 7 days in addition to thyroidectomy, however, the TRH levels exceeded that in the animals which underwent throidectomy alone. The hypothalamic TRH levels were markedly reduced in hypophysectomized rats. Conversely, in hypophysectomized rats given 7 days' thyroxine (1 and 5 microgram/100 g B.W.), the levels were increased dose-dependently. In cold-exposed rats, the plasma TSH levels roughly doubled, but the TRH levels remained unchanged. These findings strongly suggest that the feedback site of thyroxine extends not only to the pituitary gland but also to the hypothalamus, and that thyroxine has an increasing effect of the hypothalamic TRH level, though the mechanism(s) remain to be clarified.