Age-Dependent Accumulation of Hybrid Vasopressin-Oxytocin Gene Products but not Hybrid Oxytocin-Vasopressin Products in the Endoplasmic Reticulum of Brattleboro Rats

The age-dependence of the incidence of magnocellular neurosecretory neurons containing abnormal accumulations of peptide in the rough endoplasmic reticulum was examined in homozygous Brattleboro rats and in their wild-type Long Evans counterparts. Neurons in which the immunophenotype of the peptide aggregates indicate that somatic cross-over mutations involving the 5' end of the vasopressin gene and the 3' end of the oxytocin gene have occurred, increased with age in homozygous Brattleboro rats, reaching a maximum of 24 cells per hypothalamus (approximately 0.6% of the vasopressin neurons). The increase occurred in both male and female animals but was significantly greater in females. The average incidence of such cells was 6 times greater in the supraoptic than in the paraventricular nucleus. No such cells could be detected in either nucleus of Long Evans rats despite the evidence for hybrid mRNA in these animals. Moreover, no accumulation of peptide translated from the hybrid mRNAs derived from the 5' end of the oxytocin gene and the 3' end of the vasopressin gene could be detected in either Brattleboro or Long Evans animals. These results strongly suggest that the accumulation of peptide in the rough endoplasmic reticulum of vasopressin neurons in homozygous Brattleboro rats is due to an abnormality other than the somatic crossing-over mutation. A second type of abnormal magnocellular neuron with accumulations of peptide in the rough endoplasmic reticulum, in which the immunophenotype of the peptide reveals products derived only from the oxytocin precursor, was present in both Long Evans and Brattleboro rats, but did not increase with age in Brattleboro rats. The incidence of these cells was similar in the supraoptic and paraventricular nuclei.     

Cardiovascular consequences of water deprivation in female Long Evans and Brattleboro rats

Female Long Evans and Brattleboro rats were studied while water-replete and after water deprivation sufficient to cause hypovolaemia of similar degree in the two strains. A comparison was made of the blood chemistry and cardiovascular status in the two conditions, and the ability of the renin-angiotensin system, sympathoadrenal activity and (in Long Evans rats) vasopressin to influence blood pressure were assessed by pharmacological blockade of these systems. Under water-replete conditions there were significant differences between plasma variables in the two strains (Long Evans: vol., 3.67 +/- 0.07 ml/100 g b. wt., sodium, 142 +/- 0.3 mmol/l; osmolality, 290 +/- 1 mosmol/kg; Brattleboro: vol., 3.89 +/- 0.07 ml/100 g b. wt.; sodium 148 +/- 0.4 mmol/l; osmolality 304 +/- 2 mosmol/kg). Inhibition of the renin-angiotensin system (with captopril) had a slightly greater hypotensive effect in Brattleboro than in Long Evans rats. In both strains the hypotensive effects of captopril were enhanced markedly in the presence of pentolinium, and, under those conditions there was a vasopressin-dependent recovery of blood pressure in Long Evans rats that was absent in Brattleboro rats. Water deprivation caused a greater proportional reduction in body weight, and increase in plasma sodium and osmolality in Brattleboro than in Long Evans rats, although resting cardiovascular statuses were not markedly different. Despite Brattleboro rats having substantial hypernatraemia (156 +/- 1.0 mmol/l), that should have acted to inhibit renin release, they showed a profound hypotensive response to captopril that was not apparent in Long Evans rats. Thus, the absence of vasopressin in female Brattleboro rats severely affects cardiovascular adaptation to water deprivation. Comparison of the present results with published data obtained from male Long Evans and Brattleboro rats shows marked sex differences in the response to the same water deprivation protocol, and indicates that data obtained from males and females should not be cumulated.     

Metabolic alterations in the hypothalamus of the Brattleboro rat demonstrated with cytochrome oxidase histochemistry

Metabolic activity in the hypothalamus of homozygous and heterozygous Brattleboro rats and Long-Evans control rats was studied using cytochrome oxidase histochemistry. Increased metabolic activity was observed in the paraventricular nucleus (PVH), supraoptic nucleus (SON) and nucleus circularis (NC) of homozygous Brattleboro rats, and in the PVH of heterozygous rats. These results suggest that the metabolic activity of PVH and SON neurons is altered because of the inability of magnocellular neurosecretory neurons to produce vasopressin. In addition, the hyperactivity of neurons in the NC is probably related to the chronic dehydration present in these animals.     

Regulation of hypothalamic magnocellular neuropeptides and their mRNAs in the Brattleboro rat: coordinate responses to further osmotic challenge

A paradigm was developed for the chronic osmotic stimulation of homozygous diabetes insipidus rats of the Brattleboro strain, a strain that fails to synthesize vasopressin. This study examines the adaptation of 2 sets of coexisting peptide hormone magnocellular neurons in the hypothalamoneurohypophyseal system (HNS) of Long Evans (LE), Brattleboro heterozygote (HZ), and Brattleboro homozygote (DI) rats: (1) the arginine8-vasopressin (AVP)/dynorphin (DYN) neurons, and (2) the oxytocin (OT)/cholecystokinin (CCK8) neurons of the paraventricular and supraoptic nuclei, which project to the posterior pituitary. The regimen of chronic intermittent salt-loading (CISL) involved the replacement of 2% saline for normal drinking water for 18 hr/d. This protocol effectively increased plasma levels of AVP and OT in LE and HZ rats, oxytocin in DI rats, and maintained the posterior pituitary in a state depleted of AVP, OT, CCK, and peptides derived from pro-dynorphin: DYN A 1-17, DYN A 1-8, and DYN B 1-13. The ratio of pituitary DYN A 1-17 to DYN A 1-8 content in DI rats or in LE, HZ, and DI rats following 6 d of CISL suggests a preferential release of DYN A 1-17 during periods of chronic secretory activity. In response to chronic secretory activity, mRNAs for AVP, OT, DYN, and CCK increased 1.5-2-fold in all 3 AVP rat strains, with mRNAs for coexisting peptide hormones displaying parallel increases. Mutant AVP mRNA in the DI rat was expressed at very low levels and DYN mRNA in very high levels, with each of these mRNAs continuing to be regulated by CISL in a normal manner. These results suggest a regulatory relationship between AVP and OT neurons, in which vasopressin neurons are feedback-regulated by AVP, most likely via plasma osmolarity, and that oxytocin neurons are modulated by peptides derived from pro-dynorphin.     

A Role for Vasopressin in the Stress-Induced Inhibition of Gonadotrophin Secretion: Studies in the Brattleboro Rat

The effects of stress on the secretion of adrenocorticotrophin, corticosterone and luteinizing hormone (LH) in rats congenially lacking hypothalamic vasopressin (Brattleboro rats) and in normal controls of the parent strain (Long Evans) have been compared in an attempt to examine the role of vasopressin in the stress-induced depression of gonadotrophin secretion. In the Long Evans rats, stress (0.6 mg/100g histamine, ip) initiated, within 5 and 20 min respectively, significant (P <0.01, Student's t-test) increases in the plasma adrenocorticotrophin and corticosterone concentrations. It also caused a reduction in the serum LH concentration which was maximal at 5 min. By contrast, in the vasopressin deficient Brattleboro rats, stress had no effect on the serum LH concentration and produced only modest increases in pituitary adrenocortical activity compared with those in Long Evans controls. Pretreatment of both Long Evans and Brattleboro rats with dexamethasone (20μg/100 g ip, daily for 3 days) effectively abolished the pituitary-adrenal response to stress. The steroid treatment also prevented the stress-induced suppression of LH in the Long Evans rats; indeed, these animals, unlike the vehicle-treated controls, exhibited a rise in serum LH concentration within 5 min of exposure to stress. Stress did not affect the serum LH concentrations in steroid-treated Brattleboro rats. The results confirm previous reports that vasopressin is required for the full expression of the pituitary-adrenocortical response stress. They also provide novel evidence which suggests that vasopressin released in stress contributes to the impairment of gonadotrophin secretion.     

Dietary preferences of Brattleboro rats correlated with an overexpression of galanin in the hypothalamus

Galanin (GAL) is a neuropeptide cosynthesized with vasopressin (AVP) in neurons of the hypothalamo-neurohypophysial system. It increases food intake when injected into the brain and elicits an overconsumption of fat. The Brattleboro rat (DI) is genetically unable to produce AVP; the AVP-deficient-producing neurons of the hypothalamo-neurohypophysial system of DI rats are chronically stimulated and DI rats suffer from diabetes insipidus. We studied the central expression of GAL and the dietary preferences in the DI rat. GAL was overexpressed in the hypothalamus of the DI rat. GAL mRNA was higher by 1.8-fold in the supraoptic (P < 0.05) and by four-fold in the paraventricular nuclei (P < 0.001) of male and female DI rats compared with those of control Long Evans (LE) rats. However, GAL mRNA was lower in the arcuate nuclei of DI rats and equal to that of LE rats in the dorsomedian nuclei. We also measured a high preference for a lipid diet (45% of the daily consumption) when DI rats ate from a choice of the three macronutrients. Chronic infusion with deamino-8D-AVP (agonist of AVP V2 receptors) prevented the diabetes insipidus and the chronic stimulation of the hypothalamo-neurohypophysial system of the DI rats. However, the treatment did not suppress the overexpression of GAL, nor did it affect the rats' preference for a lipid diet. We conclude that the DI rat provides a novel animal model in which a spontaneous dietary preference correlates with the overexpression of one of the hypothalamic peptides, GAL.     

Chondroitin sulfate proteoglycan phosphacan/RPTPbeta in the hypothalamic magnocellular nuclei

The hypothalamo-neurohypophysial system synthesizes and releases arginine vasopressin (AVP) and oxytocin (OXT) with physiological stimulation. In the present study, we investigated localization of a chondroitin sulfate proteoglycan (CSPG), phosphacan/RPTPbeta, in the supraoptic nucleus (SON) and paraventricular nucleus (PVN) of adult rats at both the light and electron microscopic levels. Immunohistochemical analyses demonstrated stronger phosphacan/RPTPbeta immunoreactivity within the SON and PVN compared with adjacent hypothalamic areas. Double labeling experiments showed phosphacan/RPTPbeta immunoreactivity constituting punctate networks to surround the somata and dendrites of AVP- and OXT-secreting magnocellular neurons. Electron microscopic examination further revealed strong phosphacan/RPTPbeta immunoreactivity at extracellular membrane surface of some axons, somata, and dendrites of the SON, but not of synaptic junctions. Interestingly, phosphacan/RPTPbeta immunoreactivity was also observed at extracellular surface membrane between astrocytic processes and neurons rather than between magnocellular neurons. The present results indicate the high expression of the CSPG, phosphacan/RPTPbeta at the extracellular space in the hypothalamic AVP- and OXT-secreting magnocellular neurons.     

Differential expression of vasopressin alleles in the Brattleboro heterozygote

A solution hybridization/RNase protection assay for the molar quantitation of vasopressin and oxytocin mRNAs, using synthetic complementary RNA probes, is described. This assay was optimized to permit the identification of vasopressin (AVP) mRNAs containing the frame-shift point deletion causing inheritable diabetes insipidus in the Brattleboro strain of rat. Examination of RNA from hypothalamic magnocellular tissue punches found that of the 86.1 x 10(-18) mol [86.1 attomoles (amol)] of AVP mRNA detected in the Brattleboro heterozygote paraventricular (PVN) nucleus, 5.2% could be shown to be mutant AVP mRNA (AVPd RNA). The percentage of AVPd RNA increased dramatically to 18.1% after 6 d of chronic intermittent salt-loading. Similar percentages and percentage increases of AVPd RNA were detected in the heterozygote supraoptic nucleus (SON). These values were contrasted with those found in parallel studies in both Long Evans and Brattleboro homozygotes, and compared with values for oxytocin (OT) mRNA in all 3 AVP rat genotypes. The results of continued osmotic regulation of the mutant AVP gene, the low native levels of AVPd RNA found in both the Brattleboro heterozygote and homozygote, and the magnitudes of AVPd expression change with chronic osmotic challenge were interpreted as indicating that (1) in the diploid rat genome, both AVP alleles are transcribed, (2) the osmotic regulation of the mutant AVP gene is normal, and (3) the low levels of AVPd mRNA are consistent with a shorter-than-control effective mRNA half-life.     

Compensatory sprouting of uninjured magnocellular neurosecretory axons in the rat neural lobe following unilateral hypothalamic lesion

Axonal sprouting of intact neurons of the magnocellular neurosecretory system was investigated using a unilateral hypothalamic knife cut of the hypothalamoneurohypophysial tract to partially denervate the rat neural lobe (NL). Densitometric, morphometric, ultrastructural, and metabolic measures were utilized to demonstrate the compensatory response to denervation in this system. Densitometric analysis revealed a transient reduction in the intensity of vasopressin staining in the NL at 10 days postsurgery (PS) with a subsequent recovery by 20 days PS. There was a comparable initial reduction in the cross-sectional area of the NL followed by a more gradual recovery to normal by 90 days PS. Ultrastructural investigation revealed a reduction in total axon number in the NL at 10 days PS similar to the declines in vasopressin immunoreactivity and size of the NL. A subsequent partial recovery of axon number occurred, paralleling the return to normal NL size between 30 and 90 days PS. Hypertrophy of both somata and cell nuclei of magnocellular neurons in the supraoptic and paraventricular hypothalamic nuclei contralateral to the lesion was also apparent during this period. Daily measurements of urine osmolality revealed an initial transient hypoosmolality followed by a chronic hyperosmolality which persisted throughout the 90 day postsurgical period. There was a concomitant chronic decrease in both daily drinking and urine excretion volumes which began immediately following surgery. These results suggest that intact, contralateral magnocellular vasopressinergic efferents undergo compensatory sprouting as a result of partial denervation of the NL in the absence of a functional deficit in vasopressin.     

Stimulus-related changes in the dendrites of magnocellular neurones

A Golgi-Cox study was undertaken to determine whether enhanced electrical activity was associated with any morphological changes in the dendrites of the magnocellular neurones in the hypothalamic supraoptic nucleus. Brattleboro rats, animals dehydrated by administration of 2% sodium chloride solution instead of drinking water and animals given 1% sodium chloride solution and deoxycortone to induce vasopressin-dependent hypertension were compared with controls. In each of the stimulated groups, the cell bodies were hypertrophied implying that the stimuli were effective. Dendritic span (the area of a triangle drawn round, and containing the entire Golgi-stained dendritic tree) was significantly increased (p<0.01) in Brattleboro rats but was decreased by sodium chloride-induced dehydration (p<0.01). Deoxycortone treatment reversed the reduction induced by dehydration. Hippocampal cells showed no significant differences. Thus, the cells of the magnocellular system rapidly alter their morphology when stimulated but the changes are more complex than a simple hypertrophy associated with enhanced activity.     

Colocalization of vasopressin and oxytocin in hypothalamic magnocellular neurons in water-deprived rats

The posterior lobe hormones vasopressin and oxytocin are expressed in mutually-exclusive sets of magnocellular hypothamalic neurons. However, under certain functional conditions a partial coexpression has been observed. In the present study we subjected adult rats to long-term osmotic stress by water deprivation for up to 3 days.After 3 days, a marked reduction of vasopressin immunostaining was observed in the paraventricular and supraoptic nuclei as compared with controls. Coexistence of oxytocin and vasopressin occurred in a portion of the magnocellular neurons. Many fibers of the hypothalamic-neurohypophyseal tract contained both peptides. Rehydration for 24 h after 3 days of thirsting resulted in a light recovery of vasopressin immunoreactivity with almost none magnocellular neurons containing both nonapeptides. Our findings indicate that magnocellular hypothalamo neurohypophysial neurons are capable of oxytocin and vasopressin coexpression upon extended osmotic stress.     

Cardiovascular Instability Induced by Intracerebroventricular Administration of Vasopressin or Corticotrophin-Releasing Factor in Conscious Long Evans and Brattleboro Rats

Conscious Long Evans and Brattleboro rats, chronically instrumented with pulsed Doppler probes around left renal and superior mesenteric arteries and the distal abdominal aorta, had vasopressin or corticotrophin-releasing factor administered intracerebroventricularly. In Long Evans rats, vasopressin caused no cardiovascular instability, but in Brattleboro rats, vasopressin caused the development of marked oscillations in blood pressure and heart rate. Under the latter conditions, falls in blood pressure were accompanied by increases in hindquarter blood flow. Similar oscillations in blood pressure, heart rate and regional haemodynamics were seen in Long Evans and Brattleboro rats given corticotrophin-releasing factor. These results indicate that centrally-administered vasopressin and corticotrophin-releasing factor can have an important influence on the control of regional haemodynamics. In some respects the effects of the peptides resemble the changes seen during the defence reaction.     

Neuronal-glial and synaptic plasticity in the adult rat paraventricular nucleus

Using quantitative ultrastructural analysis on cells identified by immunogold postembedding immunocytochemistry, we show that magnocellular oxytocinergic neurons in the adult rat paraventricular nucleus (PVN) undergo significant neuronal-glial and synaptic changes upon stimulation. Thus, during lactation, the surface membranes of most PVN oxytocinergic somata and dendrites were directly juxtaposed; many were also contacted synaptically by the same axonal terminal ('shared' synapses). Non-oxytocinergic profiles showed few plasmalemma juxtapositions and 'shared' synapses. These ultrastructural changes are similar to those that modify oxytocin neurons in the supraoptic nucleus under the same conditions, and indicate that the whole oxytocinergic system in the hypothalamus is capable of neuronal-glial and synaptic plasticity when stimulated to release its neurohormone.     

Changes in Distribution of Cystatin C, Apolipoprotein E and Ferritin in Rat Hypothalamus after Hypophysectomy

To clarify the mechanism underlying the process of degeneration of injured CNS neurons, we have immunohistochemically examined the distribution of cystatin C, apolipoprotein E, IgG, transferrin and ferritin in the hypophysectomized rat hypothalamus. Stainings for ferritin revealed that reactive microglial cells massed in the paraventricular and supraoptic nuclei 14 days after hypophysectomy, when the degeneration of vasopressin neuronal cell bodies was apparent. Cystatin C-positive magnocellular neurons first appeared at 4 days and the number of intensely-stained cells increased rapidly up to the 7th day of hypophysectomy, followed by a decrease thereafter. Most of such cystatin C-positive neurons were simultaneously stained with anti-vasopressin serum. Accumulation of apolipoprotein E in extra-cellular spaces was obvious in the both hypothalamic magnocellular nuclei at 7 days. Several apolipoprotein E-positive cells were localized in the supraoptic nucleus, although the number of apolipoprotein E-positive cells was much smaller than that of cystatin C-positive cells. The experiments performed with the transferrin and IgG antibodies showed undetectable levels of such molecules in and around the degenerating magnocellular neurons during whole experimental periods. These findings suggest the importance of cystatin C and apolipoprotein E in the process of degeneration and/or regeneration of magnocellular neurons after hypophysectomy.     

Vasopressin-reversible increase in angiotensin-converting enzyme in specific hypothalamic nuclei of Brattleboro rats

The activity of the angiotensin-converting enzyme (ACE) (kininase II, EC 3.4.15.1) was examined in 5 discrete hypothalamic nuclei of rats lacking vasopressin (homozygous Brattleboro rats, DI, di/di) and their corresponding controls (heterozygous Brattleboro rats, HZ, di/+, and Long Evans, LE, +/+ rats), with and without hormonal replacement with arginine-vasopressin (AVP). DI rats showed a vasopressin-reversible increased ACE activity when compared with LE controls, HZ rats showing intermediate activity. These changes occurred only in the supraoptic and periventricular hypothalamic nuclei, and were absent in other hypothalamic areas studied, including the paraventricular nucleus. These results provide biochemical evidence in support of previous anatomical and physiological data, for an interaction between the brain vasopressin and angiotensin systems in discrete hypothalamic nuclei, and suggest that vasopressin could regulate the formation of brain angiotensin II by modulating the activity of the converting enzyme.     

Generation and Characterization of an Antiserum Directed Against Neurohypophyseal Prohormones

An antiserum to neurohypophyseal prohormones was generated by immunization of rabbits with a synthetic peptide fragment bridging the prohormone cleavage site between the vasopressin (AVP) and human AVP-neurophysin sequences of pro-pressophysin. Polyclonal antibodies directed against this peptide cross-reacted with intact human pro-pressophysin (ED₅₀ of 260fmol), but not with either of the final products of enzymatic processing, AVP and human AVP-neurophysin. Gel electrophoresis and Western immunoblotting of pituitary or hypothalamic extracts from multiple species including mouse, cow and man identified a protein band of molecular weight consistent with intact pro-pressophysin; in hypothalamic extracts from normally-hydrated rats no protein bands were stained, but in extracts from Brattleboro rats a faint band in the area of pro-oxyphysin was identified. Immunohistochemical studies using the antiserum demonstrated the presence of only very small amounts of immunoreactive prohormone in a few widely scattered cells in the hypothalami of normally-hydrated rats. However, after 5 days of solute loading with 2% NaCl as drinking solution, staining for intact prohormone was prominent in the supraoptic and paraventricular nuclei of the hypothalamus. Combined immunoperoxidase-immunofluorescence labeling for prohormone and either AVP-neurophysin or oxytocin-neurophysin revealed prohormone staining in both types of magnocellular neurons in rat hypothalami. These studies suggest that during states of accelerated synthesis and secretion of neurohypophyseal hormones some accumulation of intact prohormone occurs in both AVP and oxytocin magnocellular neurons.     

Comparison of the Number of Vasopressin-Producing Hypothalamic Neurons in Rats and Humans

The aim of this study was to assess the number and proportion of vasopressin-producing neurons in the hypothalamic magnocellular nuclei in rats and humans. Accurate and unbiased neuronal counts were estimated using the optical disector method. Arginine vasopressin-containing neurons were immunohistochemically visualized in formalin-fixed tissue sections. The magnocellular neurons were similar in size and morphology in both species. While the human hypothalamus contained significantly more vasopressin-containing neurons compared with the rat (36-fold increase), the proportion of vasopressin-containing neurons between species was similar. In both species, the majority of supraoptic neurons contained vasopressin, however the proportion of vasopressin-containing neurons in the human paraventricular nucleus was double that of the rat (nearly a 100-fold increase in number). These results suggest that the paraventricular nucleus contributes significantly to the release of vasopressin from the posterior pituitary in humans, whereas in rats vasopressin is mainly released by supraoptic neurons.     

Cardiac baroreflex sensitivities in conscious, unrestrained, Long Evans and Brattleboro rats

Cardiac baroreflex sensitivity (BRS; i.e. the slope of the line relating change in pulse interval to change in systemic arterial pressure) was assessed in response to an increase in mean blood pressure (MBP) evoked by methoxamine or a decrease evoked by sodium nitroprusside. Measurements were made in Long Evans (i.e. control) and in Brattleboro (i.e. vasopressin (AVP)-deficient) rats, following acute and chronic intravascular catheterization, and in water-replete and water-deprived states (the latter designed to reduce plasma volume and activate AVP-dependent mechanisms (in Long Evans rats]. There were no differences between the corresponding values for cardiac BRS in Long Evans and Brattleboro rats. Furthermore, water deprivation caused no significant changes in cardiac BRS in either strain. These results do not support the claim that absence of endogenous AVP in Brattleboro rats is associated with marked impairment of cardiac BRS, and indicate that chronic changes in circulating AVP do not cause systematic changes in cardiac BRS.     

Neuropeptide Y (NPY) and vasopressin (AVP) in the hypothalamo-neurohypophysial axis of salt-loaded or Brattleboro rats

A close anatomical relationship between nerve terminals containing neuropeptide Y (NPY) and vasopressin (AVP) has been demonstrated in the hypothalamic paraventricular (PVN) and supraoptic nuclei (SON). Furthermore, injections of NPY into the SON increased plasma concentrations of AVP in the rat. These data suggest a potential involvement of hypothalamic NPY in fluid homeostasis in the rat. Therefore, we have studied the effect of elevated plasma osmolality on the concentration of NPY and AVP in the hypothalamus and neurointermediate lobe (NIL) of the pituitary gland. Furthermore, we measured the concentration of NPY in the AVP-deficient Brattleboro rat, which suffers from diabetes insipidus and hyperosmolality. Salt-loading increased plasma osmolality and the concentration of AVP from 2.0 +/- 0.5 to 4.1 +/- 0.6 pg/ml after 7 days. The concentration of NPY in the NIL doubled after 7 days of salt-loading, from 7.9 +/- 0.6 ng/mg protein to 15.2 +/- 1.4 ng/mg protein, whereas AVP concentrations fell from 2285.7 +/- 210.9 ng/mg protein to 187.5 +/- 2.5 ng/mg protein. AVP concentrations in the ME increased transiently after 2 days of salt-loading and returned to control levels after 7 days. In contrast, NPY concentrations in the ME were unchanged at 2 days and were increased 61% after 7 days. NPY concentrations also were significantly elevated after 7 days of salt-loading in the preoptic area (POA) and mediobasal hypothalamus (MBH). The concentration of NPY in the NIL of the homozygous Brattleboro rat was 2-fold greater than in the heterozygous Brattleboro rat and 4-fold greater than in Sprague-Dawley rats used as controls.(ABSTRACT TRUNCATED AT 250 WORDS)