Regulation of KATP channel subunit gene expression by hyperglycemia in the mediobasal hypothalamus of female rats

The ATP-sensitive potassium (K(ATP)) channels are gated by intracellular adenine nucleotides coupling cell metabolism to membrane potential. Channels comprised of Kir6.2 and SUR1 subunits function in subpopulations of mediobasal hypothalamic (MBH) neurons as an essential component of a glucose-sensing mechanism in these cells, wherein uptake and metabolism of glucose leads to increase in intracellular ATP/ADP, closure of the channels, and increase in neuronal excitability. However, it is unknown whether glucose and/or insulin may also regulate the gene expression of the channel subunits in the brain. The present study investigated whether regulation of K(ATP) channel subunit gene expression might be a mechanism by which neuronal populations adapt to prolonged changes in glucose and/or insulin levels in the periphery. Ovariectomized, steroid-replaced rats were fitted with indwelling jugular catheters and infused for 48 h with saline, glucose (hyperglycemia-hyperinsulinemia), insulin and glucose (hyperinsulinemia), diazoxide (control), or glucose and diazoxide (hyperglycemia). At the end of infusions, the MBH, preoptic area, and pituitary were dissected for RNA isolation and RT-PCR. Hyperglycemia decreased Kir6.2 mRNA levels in the MBH in both the presence and absence of hyperinsulinemia. These same conditions also produced a trend toward decreased SUR1 mRNA levels in the MBH; however, it did not exceed statistical significance. Hyperglycemia increased whereas hyperinsulinemia reduced neuropeptide Y mRNA levels when these groups were compared with each other. However, neither was significantly different from values observed in saline-infused controls. In conclusion, hyperglycemia per se may alter expression of K(ATP) channels and thereby induce changes in the excitability of some MBH neurons.     

Cooperative synthesis of dopamine in rat mediobasal hypothalamus as a compensatory mechanism in hyperprolactinemia

Dopamine (DA), synthesized in the mediobasal hypothalamus by dopaminergic neurons containing two enzymes of DA synthesis–tyrosine hydroxylase and decarboxylase of aromatic L-amino acids, or by monoenzymatic non-dopaminergic neurons containing one DA synthesis enzyme in cooperation, is known to have an inhibitory effect on prolactin secretion. Deterioration of this inhibitory control leads to an increase in prolactin concentration in the blood and to the development of hyperprolactinemia syndrome. In a rat model of hyperprolactinemia induced by administration of a neurotoxin causing degeneration of dopaminergic and noradrenergic neurons, the level of DA first decreases, leading to an increase in prolactin level (decompensation stage), while later both levels are restored to normal (compensation stage). However, the mechanism of such compensation is still not clear. The aim of the present study was to analyze whether the increase in cooperative synthesis of DA by monoenzymatic neurons during hyperprolactinemia is a manifestation of a compensatory mechanism representing a particular case of neuroplasticity. The level of cooperative synthesis in the hyperprolactinemia model and in the control was estimated as the level of synthesis of DA and L-dihydroxyphenylalanine (L-DOPA)–an intermediate product of DA synthesis, when L-DOPA transfer from neurons containing tyrosine hydroxylase into neurons containing aromatic L-amino acid decarboxylase is inhibited. The level of DA synthesis during the decompensation stage was not changed, while during the compensation stage it was lower than the control. Along with a reduction in DA level, during the compensation stage an increase in the extracellular L-DOPA level in the medium was detected. Thus, the compensation of DA deficiency after degeneration of dopaminergic neurons in the mediobasal hypothalamus is due to the increase in cooperative synthesis of DA by monoenzymatic neurons containing one of the complementary enzymes of the DA synthesis pathway.     

Induction of progestin receptors in the mediobasal hypothalamus of gonadally intact male rats primed with estrogen in relation to display of lordosis behavior

The purpose of this study was to determine whether the effects of estrogen on lordosis behavior in the male rat were related to the number of progesterone (P) receptors in the mediobasal hypothalamus (MBH) and/or dependent on blood P concentration. Two groups of gonadally intact male rats were given five successive doses of 1.0 or 2.5 micrograms estradiol benzoate (EB) and tested for lordosis behavior with a male stimulus at the end of the treatment. One month later they were again injected with EB and sacrificed under the same temporal schedule, but they were not tested for lordosis so as to prevent any emotionally stressful effects of intermale cohabitation. The males given 2.5 micrograms EB more frequently displayed lordosis responses to male mounts than those receiving 1 microgram EB, with a parallel increase in the number of MBH P receptors. The total number of MBH P receptors also appeared to be higher in the animals that displayed lordosis responses (lordosis group) than in those which did not (no lordosis group). In contrast, the display of lordosis behavior was negatively correlated with blood P concentration. Comparing MBH P receptors and blood P values in the EB treated and in nonhormonally treated gonadally intact animals which had been selected for either ability or inability to spontaneously display lordosis behavior, we observed that (1) EB was capable of increasing the number of MBH P receptors in the male rat; and (2) in the absence of EB treatment blood P values were higher in the animals showing lordosis than in those which did not. These data are discussed with respect to observations made in castrated male rats and in ovariectomized females.     

Frontal deafferentation of the mediobasal hypothalamus in the neonatal rat and its effects on the preoptico-infundibular LHRH-tract

Summary By use of the peroxidase-antiperoxidase-complex (PAP) immunohistological method, the preoptico-infundibular LHRH-tract was studied in adult female rats in which frontal hypothalamic deafferentation was performed at the third or tenth postnatal day. In the former group, this LHRH-tract appeared to be similar to that of the intact controls; the animals showed regular vaginal cycles and ova were present in their oviducts. In the latter group, however, marked reduction in the number of the LHRH-nerve fibers was observed behind the sites of the deafferentation in the mediobasal hypothalamus (MBH), whereas LHRH-immunoreactive perikarya and nerve fibers containing the immunoreactive material were seen rostral to the plane of severance. In these animals reduction of LHRH-fibers in the MBH was accompanied by an anovulatory syndrome characterized by constant vaginal cornification and polyfollicular ovaries. Comparing the glial scar formation induced by the cut, significant differences were detected between the two experimental groups. In the animals deafferented on the 3rd day of life, reduction of nerve cells was seen along the cut, but LHRH-fibers crossing the thin glial scar were detectable in large numbers. On the other hand, in the animals deafferented on the 10th postnatal day, extensive glial scar tissue appeared to interrupt the LHRH-fibers rostral to the cut.     

Properties of LHRH release from a hypothalamic synaptosomal fraction of estrogen-primed ovariectomized rats

The release of luteinizing hormone-releasing hormone (LHRH) from mitochondrial-synaptosomal fractions (P₂) of basomedial hypothalamus was examined under various conditions. Less than 3% of the LHRH in P₂ suspensions was released under control conditions while the addition of 60 mM KCl or NaCl effected an 8-fold increase in LHRH as measured by radioimmunoassay. Equiosmolar sucrose effected only a 1.8-fold increase in LHRH release. The stimulatory effects of both Na⁺ and K⁺ were significantly inhibited by Mn²⁺ or La³⁺. Two forms of released LHRH were observed, one soluble and the other particulate. Soluble LHRH release was effected by hypertonic sucrose or 60 mM KCl and was not inhibited by Ca²⁺ antagonists. The release of particulate LHRH was unaffected by hypertonic sucrose, was stimulated 10-fold by 60 mM KCl, and was abolished with Ca²⁺ antagonists. These results suggest that the released soluble LHRH results from nonspecific leakage while the release of particulate LHRH reflects a Ca²⁺-dependent secretory process.     

Transmission and scanning electron-microscopic observations on tanycytes in the mediobasal hypothalamus and the median eminence of adrenalectomized rats

Summary Tanycytes in the median eminence (ME) of the rat exhibit morphological features suggesting their possible participation in transport phenomena. After adrenalectomy, which modifies the hypothalamo-hypophyseal axis, they undergo morphological changes characterized by an accumulation of lipid droplets, an increased number of bleb-like protrusions at their apex, as well as an increased pinocytosis of intraventricularly injected horseradish peroxidase (HRP). In addition, after adrenalectomy an increased number of vacuoles appears at the level of the tubero-infundibular sulci. Their intracellular location in the tanycytes is demonstrated by an intraventricular injection of HRP. The significance of these vacuoles is discussed in relation to the hydroelectrolytic modifications associated with the state of adrenalectomy.Supported by a Fonds Lekime-Ropsy AwardSupported by grants: 20472 from the Fonds de la Recherche Scientifique Médicale; and 40025 from the Fonds National de la Recherche Scientifique     

Forms of goal-directed behavior as affected by induced changes in the level of endogenous beta-endorphin in rats

Marked alterations in feeding and defense behaviour and motor activity partly resembling the effects of exogenous beta-endorphin administration were demonstrated in the experiments on rats. These alterations were observed after immunization with beta-endorphin--bovine serum albumin conjugate (two subcutaneous injections at a 7-day interval at a dose of 75 micrograms, 1 mole BSA/6 moles beta-endorphin mixed with complete Freund's adjuvant). A decrease in beta-endorphin content in some brain structures was noted. Unlike control animals, the immunized rats revealed within 3-4 weeks an increase in food intake without any rise in body weight and practically no response to handling.     

Noradrenaline turnover rate in the mediobasal and anterior hypothalamus of the rabbit

Noradrenaline turnover rate in the mediobasal and anterior hypothalamus of the rabbit. Acta Physiol. Pol., 1977, 28 (1): 39-43. The rate of noradrenaline (NA) turnover in mediobasal hypothalamus (MBH) and anterior hypothalamus (AH) of the rabbit was estimated by steady-state isotopic method with a tritiated noradrenaline (3H-NA) as a tracer. The disappearance rate of 3H-NA both in MBH and in AH was found to be biphasic; the first rapid phase of the NA half-life of about 30 min, followed by the second phase of slower decay of the half-life of 2.4 h and 10 h for MBH and AH respectively. The results suggest an existence of more than one metabolic pool of endogenous noradrenaline in MBH and AH and indicate regional difference in the metabolism of NA stores in the hypothalamus.     

Effects of estrogen and progesterone on LHRH release from a hypothalamic synaptosomal fraction of ovariectomized rats

Mitochondrial-synaptosomal fractions (P₂) from the basomedial hypothalamus of adult ovariectomized rats were employed to study the effects of estradiol benzoate (EB) and progesterone (P) on the release of luteinizing hormone-releasing hormone (LHRH). Treatment of ovariectomized rats with 5 or 50 g of EB significantly reduced the total LHRH released from P₂ under both control and K⁺-stimulated conditions. Furthermore, rats given 50 g EB demonstrated cyclic variations in the magnitude of inhibition of LHRH release. Comparison of LHRH release from P₂ of rats sacrificed at 0900 hr with that from those sacrificed at 1500 hr revealed a small persistent facilitation of LHRH release each afternoon. This facilitation, associated with an increase in the soluble component of LHRH release, was absent when rats also received 5 mg of P. No effects on LHRH release were observed when 17-estradiol alone or when P was applied to P₂ in vitro. The data show that the regulatory effects of estrogen and progesterone given in vivo on LHRH secretion can be observed in a subcellular fraction of the hypothalamus containing neurosecretory cell terminals.Supported by grants from the NIH, HD08389 and NS11753.U.S.P.H.S. Career Development Awardee, K04-HD00022     

The formation of catecholaminergic structures in the mediobasal hypothalamus of the rat

A study was made of the formation of catecholaminergic system in the medial basal hypothalamus of Wistar rat fetuses using histofluorescent method modified by de la Torre. In the periventricular region of 16-day fetuses the cells with green fluorescence characteristic for catecholamines were found. In 18-day fetuses catecholaminergic cells were found in paraventricular, arcuate and dorsomedial hypothalamic nuclei. In all the nuclei studied the number of catecholaminergic fibers and terminals of various size is greatly increased. The data obtained suggest that medial basal hypothalamus of 18-day fetuses has a complex catecholaminergic system.     

Responses to the change in the environment in pairs of male rats genetically selected for activity level

Laboratory Wistar strain rats were genetically selected for high (+A) and low (-A) activity level. In thirteen pairs of adult males of the 23rd filial generation reactions to changes in the external environment were studied. The animals were housed in breeding cages four each. Two parallel studies were conducted: in pairs simultaneously placed into a novel environment (NOV), empty cages of the same dimensions as the home cage (HC), in the second, behaviour of the second pair that remained in the HC, after removal of two cage-mates, was tested. Once a minute, for a period of one hour, the type of activity was recorded and noted whether it was an element effected in contact with the partner or without any contact. The animals +A and -A differed in the frequency of various types of activity and immobility, in the ratio between behavioural manifestations shown in or without contact as well as in the response to the type of modified environment. To changes in the situation, whether removed cage-mates from the HC or placed into NOV +A animals reacted with a high wave of environment exploration which gradually habituated. -A rats equally responded with exploration but on a lower level. In +rats we recorded more frequently exploration without contact with the partner in HC and NOV in comparison with -A, more frequent grooming, less immobility in contact and with no contact. Between +A partners there was a greater number of contacts in NOV than in HC whereas in the -A group the incidence of contact did not differ between HC and NOV. ANOVA revealed the influence of factors of genetics and environment and interaction in several behavioural categories. The simple and in time economical method demonstrated the possibility of use for the detection of differences between +A and -A lines even at relatively small changes in the external stimulatory situation.     

The bifunctional role of pro-opiomelanocortin derivatives in the mediobasal hypothalamus of the rat

In the present study, a polyclonal antibody against pro-opiomelanocortin derivatives was characterized biochemically. Its immunoreactivity with structures of the arcuate nucleus and the median eminence was investigated by means of the immunogold method and compared with its reaction on adenohypophyseal cells with and without pre-adsorption with pro-opiomelanocortin derivatives. The antiserum detects ACTH and its fragments, in particular alpha-MSH, and beta-endorphin. In the adenohypophysis gold particles are exclusively located on small secretory granules situated in the periphery of branched cells. In the perikarya of the arcuate nucleus gold particles are observed on terminal vesicles abutting from the cis-face of the Golgi apparatus, on granules in its direct vicinity and on small dense core vesicles preferentially located in the cell periphery. Immunoreactive gold-labeled fiber profiles are found in a sub- or intra-ependymal position as well as in the nuclear neuropil proper. Here axodendritic and axosomatic synapses are observed. In both situations the gold particles are mostly restricted to the small dense core vesicles and do not decorate the synaptic vesicles. In the median eminence gold labeled fibers are detected in all layers. The labeled fibers can be closely apposed to tanycytic processes, without, however, forming special contact differentiations. In direction to the perivascular layer of the external zone the labeled profiles are more frequently arranged in groups intermingled with unlabeled fibers. The axons decorated with gold particles can be freely exposed to the perivascular space or are found as single processes in close vicinity to the capillary wall. Subsequent to preincubation of the native antiserum with ACTH1-39 and ACTH18-39 (= CLIP) neither adenohypophyseal cells nor perikarya and fibers in the arcuate nucleus nor axons in the median eminence are decorated with gold particles. Preincubation of the native antiserum with alpha-MSH or beta-endorphin does not change the immunoreaction with the small, peripherally situated granules in the branched adenohypophyseal cells. In neurons of the arcuate nucleus and in fibers of the median eminence, however, the immunoreaction is completely extinguished when the antibody is pre-incubated with alpha-MSH, whereas subsequent to preincubation with beta-endorphin only the amounts of labeled structures are reduced.(ABSTRACT TRUNCATED AT 400 WORDS)     

Increased level of immunoreactive opioid peptides in the brain and adrenals of rats as affected by adaptation to physical loading

The effect of long-term (7 weeks) swimming training on the content of beta-endorphin, met- and leu-enkephalins in various brain regions and adrenal glands has been studied in Wistar rats. It has been shown that the adaptation to exercise induced an increase in the content of opioid peptides in most of the brain regions and in adrenal glands. This increase in the level of opioid peptides seems to play an important role in the increase of the resistance to stress.     

Changes in estrogen receptors in the mediobasal hypothalamus mediate the facilitory effects exerted by the male's olfactory cues and progesterone on feminine behavior in the male rat

The purpose of this study was to determine whether facilitory effects exerted by olfactory cues on lordosis behavior in the male rat involved changes in estradiol receptors at the hypothalamic level. Male rats were orchidectomized as adults. They were given either 25 micrograms estradiol benzoate (EB) alone or 25 micrograms EB and 100 micrograms progesterone (P) sequentially and exposed or not to the odor of male urine. Some of them were tested for lordosis behavior at 8 h after P. The other ones were killed 4 h after P and used for estradiol (E2) and P receptor assay in mediobasal hypothalamus (MBH). Olfactory cues were shown to increase the number of E2 receptors in both the animals given EB or EB + P. Progesterone as such appeared to be capable of increasing the number and the rate of occupancy of E2 receptors. A population of constitutive and estrogen-inducible P receptors was detected in the MBH. Since only the animals given EB + P were shown to be sensible to the facilitory effects of male urine on lordosis behavior, it may be assumed that E2 and P on one hand and olfactory cues on the other exert cumulative effects at the level of the MBH and that both a high level and a high rate of occupancy of E2 receptors are necessary for the olfactory cues to facilitate the display of lordosis behavior in the male rat.     

Effect of morphine on the concentration of monoamines in the emotiogenic zones of the hypothalamus in adult and old rats

The content of monoamines in the emotiogenic hypothalamic zones has been shown to noticeably change with aging of rats. The level of noradrenaline and serotonin increased in the ventromedial nucleus of the hypothalamus, while the concentration of noradrenaline increased in the lateral hypothalamic zone. Single i.p. injections of 10 mg/kg morphine evoked qualitatively different shifts in the monoamine concentrations in the hypothalamic emotiogenic zones of the rats of different ages: the level of dopamine increased in adult animals, while the levels of noradrenaline and serotonin dropped in old rats. It is supposed that in old age the effect of morphine on dopaminergic structures in the emotiogenic hypothalamic zones becomes more moderate, whereas that on the noradrenergic and serotonergic structures is facilitated. The age-related specificities of the morphine effect on the monoaminergic regulation of the emotiogenic hypothalamic zones can determine considerable modifications of a psychotropic effect of the drug in old age.     

Cues used by male and female hooded rats for locating a brightness change

Entries of and time spent in a novel Y-maze arm that had changed from white (during acquisition trials) to black (during retention trials) were investigated in male and female Long-Evans hooded rats after the apparatus had been horizontally rotated through 180 degrees or left undisturbed. Maze rotation reduced responsiveness to this arm in males but not in females. When each arm was associated with a different set of visual cues, males significantly chose the novel arm only in the presence of intra- and extra-maze cues either alone or in combination. Females significantly selected the novel arm only in the absence of either type of cue, and in the presence of intra-maze cues alone. However, when the duration of acquisition trials was increased from 6 to 12min, females also selected the novel arm in the presence of both intra- and extra-maze cues. It was concluded that, while female rats appeared able to use egocentric (or response-related) cues for locating the novel arm, males were more dependent on allocentric (or place-related) cues following shorter acquisition trials. Because of the importance of such cues, it seemed that the task of recognizing which maze arm had changed in brightness defined the test as one of spatial memory.     

Circadian integration of sleep-wake and feeding requires NPY receptor-expressing neurons in the mediobasal hypothalamus

Sleep and feeding rhythms are highly coordinated across the circadian cycle, but the brain sites responsible for this coordination are unknown. We examined the role of neuropeptide Y (NPY) receptor-expressing neurons in the mediobasal hypothalamus (MBH) in this process by injecting the targeted toxin, NPY-saporin (NPY-SAP), into the arcuate nucleus (Arc). NPY-SAP-lesioned rats were initially hyperphagic, became obese, exhibited sustained disruption of circadian feeding patterns, and had abnormal circadian distribution of sleep-wake patterns. Total amounts of rapid eye movement sleep (REMS) and non-REMS (NREMS) were not altered by NPY-SAP lesions, but a peak amount of REMS was permanently displaced to the dark period, and circadian variation in NREMS was eliminated. The phase reversal of REMS to the dark period by the lesion suggests that REMS timing is independently linked to the function of MBH NPY receptor-expressing neurons and is not dependent on NREMS pattern, which was altered but not phase reversed by the lesion. Sleep-wake patterns were altered in controls by restricting feeding to the light period, but were not altered in NPY-SAP rats by restricting feeding to either the light or dark period, indicating that disturbed sleep-wake patterns in lesioned rats were not secondary to changes in food intake. Sleep abnormalities persisted even after hyperphagia abated during the static phase of the lesion. Results suggest that the MBH is required for the essential task of integrating sleep-wake and feeding rhythms, a function that allows animals to accommodate changeable patterns of food availability. NPY receptor-expressing neurons are key components of this integrative function.