Differential effects of exercise on body weight gain and adiposity in obesity-prone and -resistant rats.

OBJECTIVE: To determine the effect of exercise on weight gain and adiposity in obesity-prone and -resistant rats. DESIGN: Body weight gain, fat pad weights, food intake, plasma leptin and insulin levels were assessed in outbred male Sprague-Dawley rats, which remained sedentary or were given unrestricted access to running wheels either before or after they developed diet-induced obesity (DIO) or diet-resistance (DR) on a high energy (HE; 31% fat) diet. RESULTS: When fed a low fat (4.5%) chow diet, rats which would later develop DIO (n=6) after 3 weeks on HE diet ran the same amount as DR rats (n=6). Other rats were first made DIO (n=12) or DR (n=12) after 10 weeks on HE diet and then either kept sedentary or given running wheels for 4 weeks on HE diet. DIO and DR rats ran comparable amounts but only the DIO rats reduced their body weight gain, fat pad relative to body weights and plasma leptin levels significantly, compared to their sedentary controls. Exercise had no effect on food intake in either DIO or DR rats but reduced feed efficiency (weight gain/caloric intake) in both. CONCLUSION: Although DIO and DR rats ran similar amounts, the greater reduction in body weight gain and adiposity of exercising DIO rats suggests that they are more sensitive to some metabolic or physiologic system that prevents them from increasing their intake sufficiently to compensate for their net reduction in energy stores.

The regulation of glucose-excited neurons in the hypothalamic arcuate nucleus by glucose and feeding-relevant peptides.

Glucosensing neurons in the hypothalamic arcuate nucleus (ARC) were studied using electrophysiological and immunocytochemical techniques in neonatal male Sprague-Dawley rats. We identified glucose-excited and -inhibited neurons, which increase and decrease, respectively, their action potential frequency (APF) as extracellular glucose levels increase throughout the physiological range. Glucose-inhibited neurons were found predominantly in the medial ARC, whereas glucose-excited neurons were found in the lateral ARC. ARC glucose-excited neurons in brain slices dose-dependently increased their APF and decreased their ATP-sensitive K+ channel (KATP channel) currents as extracellular glucose levels increased from 0.1 to 10 mmol/l. However, glucose sensitivity was greatest as extracellular glucose decreased to <2.5 mmol/l. The glucokinase inhibitor alloxan increases KATP single-channel currents in glucose-excited neurons in a manner similar to low glucose. Leptin did not alter the activity of ARC glucose-excited neurons. Although insulin did not affect ARC glucose-excited neurons in the presence of 2.5 mmol/l (steady-state) glucose, they were stimulated by insulin in the presence of 0.1 mmol/l glucose. Neuropeptide Y (NPY) inhibited and alpha-melanocyte-stimulating hormone stimulated ARC glucose-excited neurons. ARC glucose-excited neurons did not show pro-opiomelanocortin immunoreactivity. These data suggest that ARC glucose-excited neurons may serve an integrative role in the regulation of energy balance.

Sibutramine alters the central mechanisms regulating the defended body weight in diet-induced obese rats.

Chronic administration of sibutramine lowers body weight, presumably by altering brain monoamine metabolism. Here the effect of sibutramine on sympathoadrenal function (24-h urine norepinephrine and epinephrine levels) and arcuate nucleus (ARC) neuropeptide Y (NPY) and proopiomelanocortin (POMC) expression was assessed in diet-induced obese rats fed a low-fat diet. Chronic (10 wk) sibutramine [5 mg. kg(-1). day(-1) ip; rats fed ad libitum and injected with sibutramine (AS)] lowered body weight by 15% but only transiently (3-4 wk) reduced intake compared with vehicle-treated controls [rats fed chow ad libitum and injected with vehicle daily (AV)]. Other rats food restricted (RS) to 90% of the weight of AS rats and then given sibutramine restored their body weights to the level of AS rats when allowed libitum food intake. After reequilibration, RS rats were again energy restricted to reduce their weight to 90% of AS rats, and additional vehicle-treated rats (RV) were restricted to keep their body weights at the level of AS rats for 3 wk more. Terminally, total adipose depot weights and leptin levels paralleled body weights (AV > AS = RV > RS), although AS rats had heavier abdominal and lighter peripheral depots than RV rats of comparable body weights. Sibutramine treatment increased sympathetic activity, attenuated the increased ARC NPY, and decreased POMC mRNA levels induced by energy restriction in RV rats. Thus sibutramine lowered the defended body weight in association with compensatory changes in those central pathways involved in energy homeostasis.

Localization of glucokinase gene expression in the rat brain.

The brain contains a subpopulation of glucosensing neurons that alter their firing rate in response to elevated glucose concentrations. In pancreatic beta-cells, glucokinase (GK), the rate-limiting enzyme in glycolysis, mediates glucose-induced insulin release by regulating intracellular ATP production. A similar role for GK is proposed to underlie neuronal glucosensing. Via in situ hybridization, GK mRNA was localized to hypothalamic areas that are thought to contain relatively large populations of glucosensing neurons (the arcuate, ventromedial, dorsomedial, and paraventricular nuclei and the lateral area). GK also was found in brain areas without known glucosensing neurons (the lateral habenula, the bed nucleus stria terminalis, the inferior olive, the retrochiasmatic and medial preoptic areas, and the thalamic posterior paraventricular, interpeduncular, oculomotor, and anterior olfactory nuclei). Conversely, GK message was not found in the nucleus tractus solitarius, which contains glucosensing neurons, or in ependymal cells lining the third ventricle, where others have described its presence. In the arcuate nucleus, >75% of neuropeptide Y-positive neurons also expressed GK, and most GK+ neurons also expressed KIR6.2 (the pore-forming subunit of the ATP-sensitive K+ channel). The anatomic distribution of GK mRNA was confirmed in micropunch samples of hypothalamus via reverse transcription-polymerase chain reaction (RT-PCR). Nucleotide sequencing of the recovered PCR product indicated identity with nucleotides 1092-1411 (within exon 9 and 10) of hepatic and beta-cell GK. The specific anatomic localization of GK mRNA in hypothalamic areas known to contain glucosensing neurons and the coexpression of KIR6.2 and NPY in GK+ neurons support a role for GK as a primary determinant of glucosensing in neuropeptide neurons that integrate multiple signals relating to peripheral energy metabolism.

Presumed apoptosis and reduced arcuate nucleus neuropeptide Y and pro-opiomelanocortin mRNA in non-coma hypoglycemia.

Hypoglycemia reduces sympathoadrenal responses to subsequent hypoglycemic bouts by an unknown mechanism. To assess whether such hypoglycemia-associated autonomic failure is due to actual brain damage, male Sprague-Dawley rats underwent 1-h bouts of insulin-induced (5 U/kg i.v.) hypoglycemia (1.6-2.8 mmol/l) 1 or 3 times on alternate days. Rats remained alert and were rescued with intravenous glucose at 60-80 min. Plasma epinephrine and corticosterone responses were significantly reduced during the second and third bouts. Brains from these rats were processed by the terminal transferase-mediated deoxyuridine triphosphate-biotin nick end-labeling (TUNEL) procedure as an index of apoptotic cell death at 24, 48, or 96 h after their first bout. At 48 h, but not 24 h, TUNEL+ cells were consistently seen only in the arcuate nucleus (arcuate hypothalamic nucleus [ARC]). Hypoglycemic rats had 188% more apoptotic ARC cells (1 bout 39+/-5; 3 bouts 37+/-4) than euglycemic controls (13+/-3;P = 0.001). In situ hybridization for neuropeptide Y (NPY) and proopiomelanocortin (POMC) mRNA was performed in sections of ARC containing maximal numbers of apoptotic cells as well as in other fresh frozen brains. After 1 bout, NPY (0.041+/-0.003) and POMC (0.119+/-0.022) mRNA were decreased, respectively, by 52 and 55% vs. controls (NPY 0.076+/-0.007; POMC 0.222+/-0.020; P = 0.01). NPY (0.029+/-0.002) but not POMC (0.093+/-0.013) fell 29% further after a third bout. NPY (r = -0.721; P = 0.001) and POMC (r = -0.756; P = 0.001) mRNA levels correlated negatively with the number of apoptotic ARC cells in the same sections. Thus, non-coma hypoglycemia produces apparent apoptotic cell death with reduced NPY and POMC expression selectively in the ARC. This may contribute to the reduced counterregulatory response following repeated bouts of hypoglycemia.

Defense of body weight against chronic caloric restriction in obesity-prone and -resistant rats.

Half of Sprague-Dawley rats develop and defend diet-induced obesity (DIO) or diet resistance (DR) when fed a high-energy (HE) diet. Here, adult male rats were made DIO or DR after 10 wk on HE diet. Then half of each group was food restricted for 8 wk on chow to maintain their body weights at 90% of their respective baselines. Rate and magnitude of weight loss were comparable, but maintenance energy intake and the degree of sympathetic activity (24-h urine norepinephrine) inhibition were 17 and 29% lower, respectively, in restricted DR than DIO rats. Restricted DIO rats reduced adipose depot weights, plasma leptin, and insulin levels by 35%. Restricted DR rats reduced none of these. When fed ad libitum, both DR and DIO rats returned to the body weights of their respective chow-fed phenotype controls within 2 wk. This was associated with increased adipose mass and leptin and insulin levels only in DIO rats. Thus DR rats appear to alter primarily their lean body mass, whereas DIO rats primarily alter their adipose mass during chronic caloric restriction and refeeding.

Brain glucose sensing and body energy homeostasis: role in obesity and diabetes.

The brain has evolved mechanisms for sensing and regulating glucose metabolism. It receives neural inputs from glucosensors in the periphery but also contains neurons that directly sense changes in glucose levels by using glucose as a signal to alter their firing rate. Glucose-responsive (GR) neurons increase and glucose-sensitive (GS) decrease their firing rate when brain glucose levels rise. GR neurons use an ATP-sensitive K+ channel to regulate their firing. The mechanism regulating GS firing is less certain. Both GR and GS neurons respond to, and participate in, the changes in food intake, sympathoadrenal activity, and energy expenditure produced by extremes of hyper- and hypoglycemia. It is less certain that they respond to the small swings in plasma glucose required for the more physiological regulation of energy homeostasis. Both obesity and diabetes are associated with several alterations in brain glucose sensing. In rats with diet-induced obesity and hyperinsulinemia, GR neurons are hyporesponsive to glucose. Insulin-dependent diabetic rats also have abnormalities of GR neurons and neurotransmitter systems potentially involved in glucose sensing. Thus the challenge for the future is to define the role of brain glucose sensing in the physiological regulation of energy balance and in the pathophysiology of obesity and diabetes.

Distribution and phenotype of neurons containing the ATP-sensitive K+ channel in rat brain.

Select groups of neurons within the brain alter their firing rate when ambient glucose levels change. These glucose-responsive neurons are integrated into systems which control energy balance in the body. They contain an ATP-sensitive K+ channel (KATP) which mediates this response. KATP channels are composed of an inwardly rectifying pore-forming unit (Kir6.1 or Kir6.2) and a sulfonylurea binding site. Here, we examined the anatomical distribution and phenotype of cells containing Kir6.2 mRNA within the rat brain by combinations of in situ hybridization and immunocytochemistry. Cells containing Kir6. 2 mRNA were widely distributed throughout the brain without apparent concentration in areas known to contain specific glucose-responsive neurons. Kir6.2 mRNA was present in neurons expressing neuron-specific enolase, tyrosine hydroxylase, neuropeptide Y (NPY) and the glutamic acid decarboxylase isoform, GAD65. No astrocytes expressing glial fibrillary acidic protein or oligodendrocytes expressing carbonic anhydrase II were found to co-express Kir6.2 mRNA. Virtually all of the NPY neurons in the hypothalamic arcuate n. and catecholamine neurons in the substantia nigra, pars compacta and locus coeruleus contained Kir6.2 mRNA. Epinephrine neurons in the C2 area also expressed high levels of Kir6.2, while noradrenergic neurons in A5 and A2 areas expressed lower levels. The widespread distribution of Kir6.2 mRNA suggests that the KATP channel may serve a neuroprotective role in neurons which are not directly involved in integrating signals related to the body's energy homeostasis.

Reduced glucose-induced neuronal activation in the hypothalamus of diet-induced obese rats.

Rats predisposed to develop diet-induced obesity (DIO) preferentially activate their sympathetic nervous system during intracarotid glucose infusion [B.E. Levin, Intracarotid glucose-induced norepinephrine response and the development of diet-induced obesity, Int. J. Obesity 16 (1992) 451-457.] but their brains are generally less responsive to glucose than diet-resistant rats (DR) [B.E. Levin, K.L. Brown, A.A. Dunn-Meynell, Differential effects of diet and obesity on high and low affinity sulfonylurea binding sites in the rat brain, Brain Res. 739 (1996) 293-300.; B.E. Levin, B. Planas, Defective glucoregulation of brain alpha2-adrenoceptors in obesity-prone rats, Am. J. Physiol. 264 (1993) R305-R311.; B.E. Levin, A.C. Sullivan, Glucose-induced norepinephrine levels and obesity resistance, Am. J. Physiol. 253 (1987) R475-R481.; B.E. Levin, A.C. Sullivan, Glucose-induced sympathetic activation in obesity-prone and resistant rats, Int. J. Obesity 13 (1989) 235-246.]. Here, 1 h intracarotid glucose infusions (4 mg/kg/min) selectively increased Fos-like immunoreactivity (FLIR) in the hypothalamic paraventricular, ventromedial, dorsomedial and arcuate nuclei of inbred DR but not DIO rats. This suggests that enhanced glucose-induced sympathetic activation in DIO rats is related to a failure of glucose to produce neuronal activation in these areas.

Effect of streptozotocin-induced diabetes on rat brain sulfonylurea binding sites.

Both high and low affinity sulfonylurea receptors (SURs) reside on glucose responsive neurons where they influence cell firing and neurotransmitter release via the adenosinetriphosphate (ATP)-sensitive K+ (katp) channel. Here, the effect of diabetes on [3H] glyburide binding to SURs was assessed in male obesity-resistant Sprague-Dawley rats rendered diabetic with streptozotocin (65 mg/kg, i.p.). Additional streptozotocin-treated rats were supplemented with insulin (1.5 U/kg/ day). Streptozotocin reduced plasma insulin to 13% of control associated with hyperglycemia (25.3 +/- 1.7 mmol/l), while insulin lowered plasma glucose (9.56 +/- 1.78 mmol/l) to near control levels (7.65 +/- 0.22 mmol/l). Over 7 days, all streptozotocin-treated rats lost 12% of their initial body wt. while controls gained 1%. Despite equivalent wt. loss, streptozotocin-induced diabetes selectively increased high affinity [3H] glyburide binding in the hypothalamic dorsomedial nuclei (DMN) and ventromedial nuclei (VMN) and lateral area (LH). This was prevented by insulin injections. Low affinity binding was similarly increased in the DMN and VMN, as well as two amygdalar subnuclei but decreased in the substantia nigra, pars compacta. Insulin fully prevented these changes only in the DMN and one amygdalar nucleus and the substantia nigra. Therefore, binding to (SURs) appears to be generally upregulated in the face of hypoinsulinemia with hyperglycemia and this is prevented by insulin treatment. These and other data suggest that this combination of abnormalities in diabetes should have an adverse effect on the glucose sensing capacity of the brain.

Norepinephrine and traumatic brain injury: a possible role in post-traumatic edema.

Unilateral cerebral contusion is associated with an early (30 min) increase in norepinephrine (NE) turnover followed by a later (6-24 h) depression of turnover which is bilateral and widespread throughout the brain. Blockade of NE function during the first few hours after traumatic brain injury (TBI) impedes subsequent recovery of function without enlarging the size of the lesion. The current studies were carried out to characterize further the timing of the switch from increased to decreased NE turnover and to investigate the pathogenesis of the delayed recovery of function associated with blocking NE function. Adult male rats had unilateral somatosensory cortex contusions made with a 5 mm diameter impact piston. They were killed after 2 h and their brains analyzed for NE turnover by HPLC with electrochemical detection. In general, NE turnover (the ratio of 3-methoxy-4-hyroxyphenylglycol to NE levels) had returned to sham-lesion control levels in most brain regions by 2 h after either left or right sided contusions. The only exceptions were a persistent 87% increase at the lesion site after right-sided contusions and 22% and 32% increases in the contralateral cerebellum after right- and left-sided contusions, respectively. Blockade of alpha1-adrenoceptors by treatment with prazosin (3 mg/ kg, i.p.) 30 min prior to TBI produced edema in the striatum and hippocampus at 24 h which was not seen saline-treated rats nor in rats where NE reuptake was blocked with desmethylimipramine (DMI; 10 mg/kg, i.p.). DMI increased edema at the lesion site at 24 h, however. These data suggest that the early increase in NE release following unilateral cerebral contusion is protective and that this may act to stabilize the blood-brain barrier in areas adjacent to the injury site. Drugs that interfere with this enhanced noradrenergic function might enhance the damage caused by TBI.

Selective breeding for diet-induced obesity and resistance in Sprague-Dawley rats.

In outbred Sprague-Dawley rats, about one-half develop diet-induced obesity (DIO) on a diet relatively high in fat and energy (HE diet). The rest are diet resistant (DR), gaining weight and fat at the same rate as chow-fed controls. Here we selectively bred for high (DIO) and low (DR) weight gainers after 2 wk on HE diet. By the F5 generation, both male and female inbred DIO rats gained > 90% more weight than inbred DR rats on HE diets. Even on low-fat chow diet, DIO males were 31% and females were 22% heavier than their respective DR rats. Full metabolic characterization in male rats showed that weight-matched, chow-fed DIO-prone rats had similar energy intakes and feed efficiency [body weight (kg0.75)/energy intake (kcal)] but 44% more carcass fat than comparable DR-prone rats. Their basal plasma insulin and glucose levels in the fed state were 70 and 14% higher, respectively. But, when fasted, DIO-prone oral glucose tolerance results were comparable to DR-prone rats. Chow-fed DIO-prone males also had 42% greater 24-h urine norepinephrine levels than DR-prone males. During 2 wk on HE diet, DIO rats ate 25% more, gained 115% more weight, had 36% more carcass fat, and were 42% more feed efficient than comparable DR rats. Fasted HE diet-fed DIO rats developed frank glucose intolerance during a glucose tolerance test with 55 and 158% greater insulin and glucose areas under the curve, respectively. Thus the DIO and DR traits in the outbred Sprague-Dawley population appear to be due to a polygenic pattern of inheritance.

Histological markers of neuronal, axonal and astrocytic changes after lateral rigid impact traumatic brain injury.

The model of lateral, rigid impact traumatic brain injury is widely used but remains relatively poorly characterized by comparison with fluid percussion injury models. Thus, whilst the gross morphological changes that occur over the short- and long-term post-injury have been described, more subtle measures of neuronal injury and activation, and markers of axonal and glial reactions have not been investigated, complicating interpretation of data from this model. To address this issue, a variety of neurohistological markers were examined in adult male rats which had been subjected to open brain, lateral rigid impact injury. A piston device was unilaterally driven 3.0 mm into the somatosensory cortex at a speed of 3.2 m/s. Neuronal activation evidenced by Fos-like immunoreactivity showed a complex pattern at 3 h after injury which appeared to be related both to proximity to the impact site and cortical efferent connectivity. At 24 h after injury, acid fuchsin staining demonstrated dying neurons in the margin of the injury and in ipsilateral hippocampus and dorsal thalamus. Injured cells identified by heat-shock protein immunoreactivity showed a similar distribution. Axonal injury demonstrated with 68 kDa neurofilament immunoreactivity was more widely distributed. Less axonal damage was found with increasing distance from the injury site. At 7 days post-injury, glial fibrillary acidic protein immunoreactive astrocytes were prolific in the ipsilateral thalamus, hippocampus and striatum and throughout the injured cortex. In general, controlled, lateral rigid impact injury provides a more focused injury than is seen with lateral fluid percussion which may have implications for the behavioral deficits seen in this injury model.

Location and effect of obesity on putative anorectic binding sites in the rat brain.

Anorectic drugs such as mazindol bind to a class of low-affinity, sodium-sensitive sites in the brain which are affected by ambient glucose concentrations and a predisposition to develop diet-induced obesity (DIO). This study used quantitative autoradiography of 10 nM 3H-mazindol binding to identify the cellular location of these putative anorectic binding sites in the brain and to assess the way in which the development of DIO affected their binding. We previously showed that chow-fed, obesity-prone rats have widespread increases in brain 3H-mazindol binding to these low-affinity sites as compared with diet-resistant (DR) rats. Here, low-affinity 3H-mazindol binding was assessed in the brains of eight rats which developed DIO vs. eight which were DR after three months on a high-energy diet. DIO rats gained 89% more weight and had 117% higher plasma insulin levels but no difference in plasma glucose levels compared with DR rats. Along with these differences, low-affinity 3H-mazindol binding in DIO rats was identical to that in DR rats in all of the 23 brain areas assessed. This suggested that this binding was downregulated by the development of obesity in DIO rats. In other chow-fed rats, stereotaxic injections of 5,7-dihydroxytryptamine and 6-hydroxydopamine (6OHDA) to ablate serotonin and catecholamine nerve terminals in the ventromedial nucleus of the hypothalamus (VMN) had no effect on 3H-mazindol binding. However, ibotenic acid injected into the VMN, substantia nigra, pars reticulata, and pars compacta destroyed intrinsic neurons and/or their local processes and decreased low-affinity 3H-mazindol binding by 13%-22%. Destruction of dopamine neurons in the substantia nigra, pars compacta, and noradrenergic neurons in the locus ceruleus with 6OHDA also reduced 3H-mazindol binding in those areas by 9% and 12%, respectively. This suggested that up to 22% of putative anorectic binding sites may be located on the cell bodies of dopamine, norepinephrine, and other neurons, but not on serotonin or catecholamine nerve terminals in the brain. Binding to these sites may be downregulated by the development of DIO, possibly as a result of the concomitant hyperinsulinemia.

Dysregulation of arcuate nucleus preproneuropeptide Y mRNA in diet-induced obese rats.

Neuropeptide Y (NPY) neurons in the hypothalamic arcuate nucleus (ARC) produce metabolic and physiological effects that promote the development and maintenance of obesity. In turn, NPY metabolism in these neurons is inhibited by dopamine release. In this study, ARC prepro-NPY mRNA and ARC/median eminence (ME) dopamine turnover were assessed in chow-fed male Sprague-Dawley rats prone to develop diet-induced obesity (DIO) or to be diet resistant (DR) when fed a high-energy (HE) diet. By in situ hybridization, DIO-prone rats had 39% more ARC NPY mRNA expression than DR-prone rats under chow-fed conditions. DIO-prone rat ARC/ME dopamine levels were 14% higher, but dopamine half-life was 176% longer and turnover was 59% less than DR-prone rats. Neither a 48-h fast nor 50% energy intake restriction for 5 days affected the already increased ARC NPY mRNA levels in DIO-prone rats. Both manipulations increased NPY expression to the level of DIO-prone rats in DR-prone rats by 23 and 35%, respectively. Finally, when fed HE diet for 2 wk, neither DIO- nor DR-prone rats altered their ARC NPY expression despite the development of obesity and hyperinsulinemia in DIO rats. Thus DIO-prone rats overexpress and fail to regulate ARC NPY mRNA to energy restriction or hyperinsulinemia. This dysregulation is possibly secondary to reduced inhibition because of defective ARC/ME dopamine turnover. Both may be important predisposing factors in the development of DIO.

Intracarotid glucose selectively increases Fos-like immunoreactivity in paraventricular, ventromedial and dorsomedial nuclei neurons.

Perfusion of the forebrain with glucose at concentrations which alter neither plasma insulin nor glucose levels leads to sympathetic activation in some rats. We used the expression of Fos-like immunoreactivity (FLI) as an index of neuronal activation to examine the anatomic substrate underlying this phenomenon. Male Sprague-Dawley rats were infused via the right internal carotid artery with glucose (4 mg/kg/min) or equiosmolar mannitol for 60 min. They were killed 3 h after infusion onset and their brains reacted for FLI. As compared to mannitol-infused controls, 105% and 117% more neurons in hypothalamic ventromedial nucleus (VMN) and parvocellular portion of the paraventricular nuclei (PVN) of glucose-infused rats showed FLI, respectively. Importantly, only about half the glucose-infused rats showed increased FLI cells in these areas when compared to controls. In these same animals, glucose also significantly activated cells in the dorsomedial n. There was little FLI expressed in the magnocellular neurons of the PVN. This selective glucose response was bilateral in keeping with the bilateral distribution of India ink to midline hypothalamic structures following unilateral carotid infusions. Retrograde transport of cholera toxin B from medullary and thoracic spinal cord sympathetic outflow areas showed labeling of about 10% of PVN neurons with FLI activated by intracarotid glucose. There was no double labeling of VMN neurons. This supports the presence of anatomic pathways by which a subpopulation of glucose responsive PVN neurons might activate the sympathetic outflow areas in the medulla and spinal cord. The apparent bimodal distribution of glucose-induced activation of VMN and PVN neurons is in keeping with a similar bimodal pattern of sympathetic activation which obesity-prone but not obesity-resistant rats show following glucose infusions. Taken together, these data support a role for glucose-sensitive VMN and parvocellular PVN neurons in the weight gain phenotype specific sympathetic activation to glucose.

Low-affinity sulfonylurea binding sites reside on neuronal cell bodies in the brain.

The antidiabetic sulfonylurea drugs bind to sites associated with an ATP-sensitive potassium (Katp) channel on cell bodies and terminals of neurons which increase their firing rates or transmitter release when glucose concentrations rise or sulfonylureas are present. High-affinity sulfonylurea binding sites are concentrated in areas such as the substantia nigra (SN) where glucose and sulfonylureas increase transmitter release from GABA neurons. But there is a paucity of high-affinity sites in areas such as the hypothalamic ventromedial nucleus (VMN) where many neurons increase their activity when glucose rises. Here we assessed both high- and low--affinity sulfonylurea binding autoradiographically with 20 nM [3H]glyburide in the presence of absence of Gpp(NH)p. Neurotoxin lesions with 6-hydroxydopamine (6-OHDA), 5,7-dihydroxytryptamine (5,7-DHT) and ibotenic acid were used to elucidate the cellular location of the two sites in the VMN, SN and locus coeruleus (LC). In the VMN, 25% of the sites were of low affinity. Neither 6-OHDA nor 5,7-DHT affected [3H]glyburide binding, while ibotenic acid reduced the number of VMN neurons and abolished low-affinity without changing high-affinity binding. In cell-attached patches of isolated VMN neurons, both 10 mM glucose and 100 microM glyburide decreased the open probability of the Katp channel suggesting that the low-affinity binding site resides on these neurons. In the SN pars reticulata, ibotenic acid reduced the number of neurons and high-affinity [3H]glyburide binding was decreased by 20%, while 6-OHDA had no effect. In the SN pars compacta, both 6-OHDA and ibotenic acid destroyed endogenous dopamine neurons and selectivity ablated low-affinity binding. In the LC, 6-OHDA destroyed norepinephrine neurons and abolished low-affinity binding. These data suggest that low-affinity sulfonylurea binding sites reside on cell bodies on VMN, SN dopamine and LC norepinephrine neuron cell bodies and that high-affinity sites may be on axon terminals of GABA neurons in the SN.

Alpha 1-adrenoceptor blockade increases behavioral deficits in traumatic brain injury.

Experimental enhancement of noradrenergic activity following traumatic brain injury (TBI) accelerates behavioral recovery if performed at a time when brain norepinephrine (NE) turnover is decreased. But, since NE turnover is markedely increased immediately after TBI, the present study was undertaken to evaluate the effect of modulating these early changes in NE metabolism on recovery of function. Rats were pretrained on a modified beam walking task. Thirty minutes prior to unilateral somatosensory cortex contusion they were treated with a NE reuptake blocker [desmethy-limipramine (DMI); 10 mg/kg, ip, n = 6] or an alpha 1-adrenoreceptor antagonist [prazosin (PRZ); 3 mg/kg, ip, n = 6]. PRZ pretreatment markedly worsened beam walking performance throughout the 3 weeks following injury, whilst DMI pretreatment did not affect performance compared to injured controls (n = 4). Despite the marked behavioral deficits, PRZ-treated animals showed no apparent worsening of histological damage (n = 11 per group) and lesion size was the same in all groups. In separate experiments (n = 4 per group), PRZ lowered basal blood pressure and prevented the rise in pressure immediately following TBI. However, blood pressures in the three groups came to the same level within 20 sec following TBI. This suggest that the action of PRZ was not simply due to hypotension-induced ischemia. It is possible that blockade of alpha 1-adrenoreceptors in the immediate posttrauma period leads to enhancement of excitatory neurotransmission, which exacerbates behavioral deficits.

In vivo and in vitro regulation of [3H]glyburide binding to brain sulfonylurea receptors in obesity-prone and resistant rats by glucose.

Select brain neurons increase their firing rate when ambient glucose levels rise, possibly via a neuronal ATP-sensitive K+ (KATP) channel and its associated sulfonylurea receptor (SUR). We used receptor autoradiographic binding of 20 nM [3H]glyburide (in the presence or absence of Gpp(NH)p which blocks binding to low-affinity sites) to assess the in vivo and in vitro effects of altering glucose availability upon high- and low-affinity binding to brain SUR. Since the brain's ability to monitor and regulate glucose metabolism is critical to maintenance of energy balance, testing was done in chow-fed male Sprague-Dawley rats which had an underlying predisposition to develop either diet-induced obesity (DIO-prone) or to be diet-resistant (DR-prone) when subsequently fed a high-energy diet. Under control conditions, both in vivo and in vitro studies showed DIO-prone rats to have reduced levels of low-, but not high-affinity [3H]glyburide binding in most forebrain areas. As compared to equiosmolar infusions of mannitol, 60 min unilateral intracarotid glucose infusions at 4 mg/kg/min in awake rats reduced low-affinity [3H]glyburide binding in numerous hypothalamic and amygdalar areas of both DR- and DIO-prone rats with little effect on high-affinity binding. Only in the paraventricular nucleus of DR-prone rats was there a phenotype-specific downregulation of low-affinity binding. Brain sections from other rats were incubated with [3H]glyburide in the presence of 0, 5 or 10 mM glucose. The resultant in vitro effects of glucose were more variable and widespread than intracarotid infusions. Here, glucose often increased low-affinity [3H]glyburide binding, particularly in DR-prone rats at 5 mM. Again, there was little effect on high-affinity binding. Thus, glucose may affect the firing of glucose-responsive neurons by indirectly altering KATP channel function via its effects on low-affinity cell body SUR.