Pauline E. McKibbin

Acute fenfluramine administration reduces neuropeptide Y concentrations in specific hypothalamic regions of the rat: possible implications for the anorectic effect of fenfluramine.

Neuropeptide Y (NPY), a powerful central appetite stimulant, coexists in several hypothalamic areas with serotonin, which suppresses feeding. This study investigated the effect of acute administration of the serotonergic drug, fenfluramine, on NPY concentrations in microdissected hypothalamic nuclei. Adult male Wistar rats were given fenfluramine (10 mg/kg, n = 7) or saline (n = 7), intraperitoneally 1 h before darkness. Food was presented immediately before darkness and the rats were killed during the first 4 h of darkness. Fenfluramine injection significantly reduced food intake. In fenfluramine-injected animals, NPY levels in the ventromedial and dorsomedial nuclei and in the lateral hypothalamic and lateral preoptic areas were significantly lower than in saline-injected controls. The ventromedial and dorsomedial nuclei and the lateral hypothalamic area are sites which mediate the hyperphagic action of centrally injected NPY. Selective NPY changes in specific nuclei after fenfluramine injection suggest functional interaction between NPYergic and serotonergic systems, and may indicate that NPY is involved in mediating the anorectic effect of serotonergic agents.

Insulin deficiency is a specific stimulus to hypothalamic neuropeptide Y: A comparison of the effects of insulin replacement and food restriction in streptozocin-diabetic rats ☆

Untreated insulin-deficient diabetes causes hyperphagia and neuroendocrine disturbances that may be partly mediated by increased hypothalamic activity of neuropeptide Y (NPY), a potent central appetite stimulant. The metabolic signal that stimulates hypothalamic NPY is unknown. This study aimed to determine whether insulin deficiency or hyperglycemia was responsible. Regional hypothalamic NPY concentrations were compared in streptozocin-diabetic (STZ-D) rats rendered nearly normoglycemic by either insulin replacement or food restriction. Untreated STZ-D rats were hyperphagic and showed significantly increased (p less than 0.01) hypothalamic NPY concentrations in the arcuate nucleus and lateral hypothalamic area. Once-daily ultralente insulin injections corrected hypoinsulinemia and hyperglycemia, abolished hyperphagia, and normalized NPY concentrations in all hypothalamic regions. By contrast, food restriction effectively lowered glycemia without raising insulin levels. In these underfed diabetic rats, NPY concentrations rose further and were significantly higher than nondiabetic and untreated diabetic levels in most hypothalamic regions. We conclude that insulin deficiency is a major stimulus to hypothalamic NPY in STZ-D, whereas hyperglycemia may exert an inhibitory influence. These findings support the hypothesis that hypothalamic NPY responds to specific metabolic cues and is involved in regulating energy balance and conserving body weight.

Hypothalamic neuropeptide Y receptor characteristics and NPY-induced feeding responses in lean and obese Zucker rats

Increased hypothalamic neuropeptide Y levels have previously been demonstrated in several hypothalamic nuclei of the (fa/fa) Zucker rat. This study set out to characterise hypothalamic NPY receptors in both genotypres and to study the effect of exogenous NPY on feeding behavior in these rats. Spontaneous daytime food intake was raised in the obese rat (p less than 0.05). Total hypothalamic receptor density (Bmax) was reduced in the obese rat compared with the lean rat (by 56%, p less than 0.005), but affinity remained unaltered. The lowest dose of NPY tested (23.5 pmol) stimulated daytime feeding in lean rats after 1, 2 and 3 hours but was inaffective in the obese rat (p less than 0.05). At two higher doses (235 pmol and 2.35 nmol), NPY was equipotent in both genotypes over 1 and 2 hours but NPY-induced feeding was attenuated over 3 hours in the obese rat. These results suggest an overactive endogenous NPYergic system in the obese (fa/fa) rat which might contribute to hyperphagia and obesity in this strain.

Hypothalamic neuropeptide Y disturbances in the obese (cp/cp) JCR:LA corpulent rat

Regional hypothalamic neuropeptide Y (NPY) concentrations were compared between cp/cp JCR:LA corpulent rats, which were grossly obese, hyperphagic, and hyperinsulinemic, and lean (+/+) controls. In freely fed cp/cp rats, NPY levels in the arcuate nucleus (ARC) were 31% higher than in lean rats (p less than 0.001). In lean rats, chronic food restriction significantly raised NPY levels by 22% in the ARC (p less than 0.05) and by 44% in the dorsomedial nucleus (DMH; p less than 0.05). By contrast, food-restricted cp/cp rats showed no change in the ARC, but NPY levels rose in the DMH (by 36%; p less than 0.05) and ventromedial nucleus (31%; p less than 0.05). Increased NPY levels in the ARC, the major site of hypothalamic NPY synthesis, suggests increased NPYergic activity in cp/cp rats; given the central actions of NPY, this could contribute to hyperphagia, obesity, and hyperinsulinemia in this syndrome. Abnormal NPY responses to food deprivation further suggest dysregulation of NPY in cp/cp rats.

Peripheral insulin administration attenuates the increase in neuropeptide Y concentrations in the hypothalamic arcuate nucleus of fasted rats

Fasting increases neuropeptide Y (NPY) concentrations in the arcuate nucleus (ARC), its site of synthesis, and in other regions of the rat hypothalamus. Neuropeptide Y is a potent central orexigenic agent and may therefore stimulate appetite during fasting. We tested the hypothesis that low plasma insulin levels stimulate ARC levels of NPY in fasted rats. Compared with freely fed controls (n = 8), rats fasted for 72 h (n = 8) showed significantly lower plasma insulin levels (28.9 +/- 1.6 vs. 52.6 +/- 5.7 pmol/l; p < 0.001) and higher ARC NPY concentrations (14.2 +/- 1.8 vs. 8.4 +/- 2.2 fmol/micrograms protein; p < 0.001). Fasted rats treated with subcutaneous insulin (5 U/kg/day; n = 10), which nearly normalized plasma insulin (46.6 +/- 2.8 pmol/l), showed intermediate ARC NPY levels (11.2 +/- 1.4 fmol/micrograms protein; p < 0.01 vs. controls and untreated fasted rats). Insulin administered peripherally, therefore, attenuates fasting-induced NPY increases in the ARC, supporting the hypothesis that hypoinsulinemia stimulates hypothalamic NPY.

Increased neuropeptide Y concentrations in the lateral hypothalamic area of the rat after the onset of darkness: Possible relevance to the circadian periodicity of feeding behavior

Neuropeptide Y (NPY) is a major hypothalamic peptide which powerfully stimulates feeding when injected into the hypothalamus and is implicated in circadian rhythmicity. To investigate whether NPY is involved in the increased feeding that follows the onset of darkness in rats, NPY levels were measured in discrete hypothalamic areas before and after darkness. Four groups of eight adult female Wistar rats were habituated to a 12:12 hour light:dark cycle, with food presented at the onset of darkness (19.00 hours). One group was sacrificed during the 3 hours before darkness and another in the first 2.5 hours after darkness, with food provided as usual. To distinguish any effects of feeding itself, the study was repeated with two further groups, but food was not provided after darkness. Seven hypothalamic regions were microdissected from slices of fresh brain and acid-extracted for radioimmunoassay of NPY. NPY levels (fmol/microgram protein) were significantly higher (p less than 0.01) in the lateral hypothalamic area (LHA) of the dark-phase group in both studies. In the other six regions, NPY levels did not differ between light and dark phases. The LHA regulates the circadian rhythmicity of feeding and NPY injection here stimulates feeding. Alterations in NPY in the LHA around the onset of darkness may be related to the initiation of dark-phase feeding.

The effect of dexamethasone on neuropeptide y concentrations in specific hypothalamic regions

Neuropeptide Y (NPY) is a major hypothalamic peptide which is implicated in the regulation of energy balance and in the activation of the hypothalamo-pituitary adrenal axis. This study aimed primarily to determine the effects on regional hypothalamic NPY levels, of catabolism and weight loss induced in rats by the synthetic glucocorticoid, dexamethasone, injected daily at a dose of 0.4 mg/kg for 7 days. NPY concentrations were significantly raised in the paraventricular nucleus (PVN) of male Wistar rats (45%, p = 0.009; n = 10) compared with saline-injected controls (n = 10). Body weight (p less than 0.001) and food intake (p less than 0.001) were significantly reduced, plasma insulin concentrations were increased (p less than 0.001), but there was no change in glucose concentrations. Chronic dexamethasone treatment did not cause the marked NPY increases in the arcuate nucleus (ARC) and other hypothalamic regions which have been observed in other catabolic states causing weight loss. One possible explanation is the high insulin levels induced by dexamethasone, which may have prevented compensatory hyperphagia by suppressing an increase in hypothalamic NPYergic activity. We also examined the acute effects of a single dexamethasone injection on regional hypothalamic levels, to determine whether the drug had a direct action separate from that due to sustained weight loss. In the acute study, groups of rats (n = 7) were examined at 4 h after a single injection of dexamethasone or saline. NPY concentrations were significantly increased in the lateral hypothalamic area (LHA), (60%, p = 0.008) when compared with saline-injected controls, but there was no change in body weight or glucose or insulin concentrations during the 4h interval. Altered transport or release of NPY in the lateral hypothalamic area may be a result of acute feedback regulation by glucocorticoids on the hypothalamus.

Unchanged hypothalamic neuropeptide Y concentrations in hyperphagic, hypoglycemic rats: Evidence for specific metabolic regulation of hypothalamic NPY

Hypothalamic concentrations of neuropeptide Y (NPY), a potent central appetite stimulant, increase dramatically in food-restricted and insulin-deficient diabetic rats. This suggest that NPY may drive hyperphagia in these conditions, which are characterized by weight loss and insulin deficiency. To test the hypothesis that insulin deficiency and weight loss are specific stimuli to hypothalamic NPY, we measured NPY concentrations in individual hypothalamic regions in rats with hyperphagia caused by insulin-induced hypoglycemia. Groups of 8 male Wistar rats were injected with ultralente insulin (20-60 U/kg) to induce either acute hypoglycemia (7 h after a single injection) or chronic hypoglycemia (8 days with daily injections). In hypoglycemic rats, plasma insulin concentrations were increased 6- to 7-fold compared with saline-injected controls; food intake was significantly increased with acute and chronic hypoglycemia and weight gain was significantly increased in the chronically hypoglycemic group. NPY concentrations were measured by radioimmunoassay in 8 hypothalamic regions microdissected from fresh brain slices. NPY concentrations were not increased in any region in either acute or chronic hypoglycemia. NPY therefore seems unlikely to mediate hyperphagia in hyperinsulinemia-induced hypoglycemia, supporting the hypothesis that weight loss is a specific stimulus to hypothalamic NPY and that insulin deficiency may be the metabolic signal responsible.

Prolonged pulsatile release of gonadotropin-releasing hormone from the guinea pig hypothalamus in vitro

The sexually mature mammal secretes luteinizing hormone in a pulsatile fashion. This is presumed to depend on the intermittent release of hypothalamic gonadotropin- releasing hormone (GnRH). The isolated guinea pig hypothalamus has been studied because, in this species, as in primates, the pulse generator appears to reside within the medial basal hypothalamus. The basal 2 mm of guinea pig hypothalami were rapidly removed and perifused at 37 degrees C with Krebs-Ringer solution containing 20 mM bacitracin gassed with 95% O2, 5% CO2. The eluates were sampled at 15 and 5 min intervals and pulsatile patterns of GnRH were consistently observed for periods up to 72 h. There was no difference in GnRH levels from hypothalami of intact and ovariectomized animals. Simultaneous measurement of TRH and somatostatin disclosed independent pulses of both neurohormones which did not coincide with GnRH, indicating that the peaks were secretory episodes not artefacts generated by varying perifusion rates. The hypothalami disclosed no histologic evidence of necrosis when examined after 20 h perifusion.

Acute hyperinsulinaemia causes an increase in neuropeptide Y (NPY) concentrations in the hypothalamic arcuate nucleus of the rat

NPY, a major hypothalamic peptide, stimulates feeding, insulin secretion and weight gain when injected intrahypothalamically. NPY synthesis in the arcuate nucleus (ARC) and levels in NPY-sensitive hypothalamic regions rise in diabetes and starvation, possibly stimulated by insulin deficiency. High pharmacological intracerebroventricular insulin levels inhibit NPY synthesis in the ARC, but the effects of hyperinsulinaemia have not been studied. We therefore measured regional hypothalamic NPY levels in rats during a hyperinsulinaemic, euglycaemic clamps. Male Wistar rats with implanted jugular cannulae, fasted for 24 hrs, were infused with insulin (128.2 mU1⁻¹ at 780 μl hr⁻¹ (n = 7) together with variable-rate glucose to maintain euglycaemia (3.9± 0.1 mmoll⁻¹), or saline (n=8; glycaemia, 4.0±0.5 mmoll⁻¹), for 2.5 hrs. Insulin levels were 80.2±10.4 mUl⁻¹ in insulin-infused rats and 16.7±11.7 mUl⁻¹ in saline-treated controls (p<0.001). NPY levels were measured by radioimmunoassay in the ARC and 7 other hypothalamic regions. NPY levels in the ARC were significantly higher in hyperinsulinaemic than in control rats (4.80±1.17 vs 2.54±0.58 fmol μg protein⁻¹; p<0.001), but comparable with controls in all other regions. Acute physiological hyperinsulinaemia therefore increase NPY levels selectively in the ARC. Insulin may cause NPY accumulation in the ARC by blocking its transport to NPY-sensitive areas, consistent with the suggested inhibition by insulin of NPY and the postulated role of insulin as a satiety factor.