RESUMO
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.
Assuntos
Apoptose , Núcleo Arqueado do Hipotálamo/metabolismo , Hipoglicemia/fisiopatologia , Neuropeptídeo Y/genética , Pró-Opiomelanocortina/genética , RNA Mensageiro/antagonistas & inibidores , Animais , Encéfalo/patologia , Encéfalo/fisiopatologia , Hipoglicemia/induzido quimicamente , Hipoglicemia/metabolismo , Hipoglicemia/patologia , Hipoglicemiantes , Hibridização In Situ , Marcação In Situ das Extremidades Cortadas , Insulina , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
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.
Assuntos
Encéfalo/fisiologia , Expressão Gênica/fisiologia , Glucoquinase/genética , Canais de Potássio Corretores do Fluxo de Internalização , Animais , Núcleo Arqueado do Hipotálamo/metabolismo , Sequência de Bases/genética , Encéfalo/metabolismo , Hipotálamo/metabolismo , Hibridização In Situ , Masculino , Canais de Potássio/metabolismo , Punções , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Distribuição TecidualRESUMO
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.
Assuntos
Aloxano/farmacologia , Núcleo Arqueado do Hipotálamo/fisiologia , Glucose/farmacologia , Neurônios/fisiologia , Animais , Núcleo Arqueado do Hipotálamo/efeitos dos fármacos , Técnicas In Vitro , Masculino , Potenciais da Membrana/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Técnicas de Patch-Clamp , Ratos , Ratos Sprague-Dawley , Tolbutamida/farmacologiaRESUMO
The projection of the nucleus isthmi to the ipsilateral optic tectum was examined in normal goldfish. This was compared to the projection in animals in which the entire visual field had been induced to compress onto a rostral half tectum by caudal tectal ablation. The isthmo-tectal projection was examined by making localized injections of horseradish peroxidase into the optic tecta and observing the patterns of labeled cells within the nucleus isthmi. The teleost nucleus isthmi consists of a cell sparse medulla covered by a cellular cortex, which is thick on the rostral, medial, and dorsal surfaces of the nucleus. Almost all isthmic cells projecting to the tectum were located in the area of thick cortex. In normal fish, rostral tectal injections labeled cells in the rostroventral portion of the thick cortex; injections midway in the rostrocaudal tectal axis labeled more caudodorsally located cells, and caudal tectal injections labeled cells a little further caudally in extreme dorsal cortex. The rostroventral to caudodorsal isthmic axis was therefore seen to project rostrocaudally along the tectum. This topography contrasts somewhat with the situation seen in amphibia where the rostrocaudal tectal axis receives projections from the rostrocaudal isthmic axis. In fish with half-tectal ablations, injections near the caudal edge of the half tectum (at a site that had originally been midtectal) labeled cells that had previously projected to caudal tectum. Rostral tectal injections in fish with compression of the visual field gave a normal pattern of labeled isthmic cells. The results indicate that a topographically ordered isthmo-tectal projection exists in goldfish that may be induced to compress onto a half tectum.
Assuntos
Cyprinidae/anatomia & histologia , Carpa Dourada/anatomia & histologia , Colículos Superiores/anatomia & histologia , Tegmento Mesencefálico/anatomia & histologia , Animais , Mapeamento Encefálico , Plasticidade Neuronal , Colículos Superiores/patologia , Tegmento Mesencefálico/patologia , Vias Visuais/anatomia & histologiaRESUMO
In channel catfish the ganglion cell axons leave the retina via a ring of approximately 13 separate optic papillae. Each papilla serves an area of retina extending from the central zone of the retina to the periphery. Papillae located at a dorsal position in the ring serve exclusively dorsal retina. Ventrally located papillae, however, have an exaggerated peripheral retinal representation, so that they serve mostly ventral retina but also some areas of peripheral retina dorsal to the nasal and temporal poles. The ganglion cell axon bundles departing from the retina via individual papillae were labelled with horseradish peroxidase, and sections of the optic pathway were examined to reveal the topographic organization of the fibers. The topographic order of the optic nerve was dissimilar to that of cichlids and goldfish. Fibers from individual papillae remained together throughout the optic nerve. Close to the optic nerve head, the papillae were arranged as a continuum around the U-shaped optic nerve, without the discontinuity in the representation of the ventral retina seen in other fish. Fibers associated with the dorsal papillae were located at the tip of the caudolateral arm of the U, and fibers from ventral papillae were on the rostromedial arm. Fibers from nasally and temporally located papillae were found on the base of the U. By the level of the optic chiasm the U shape had flattened out but retained the relative ordering of the papillae. Rotation of the nerve as it became the optic tract brought the representation of the ventral papillae to the dorsal pole of the tract, and the dorsal papillae to the ventral tract. It was only in the optic tract that rearrangement of fibers became apparent. As described above, the axons of some ganglion cells in dorsal, peripheral retina left the retina and travelled through the optic nerve with axons from extreme ventral retina. In the optic tract, these dorsal fibers joined the main body of fibers from the dorsal retina. The significance of these observations for theories of fiber rearrangement is discussed.
Assuntos
Peixes-Gato/anatomia & histologia , Ictaluridae/anatomia & histologia , Fibras Nervosas/citologia , Nervo Óptico/anatomia & histologia , Retina/citologia , Células Ganglionares da Retina/citologia , Vias Visuais/anatomia & histologia , Animais , Quiasma Óptico/anatomia & histologiaRESUMO
The retinal organization associated with the multiple optic papillae of the catfish Ictalurus punctatus was examined. In each retina from ten to 17 papillae form an oval ring (which is wider dorsoventrally than mediolaterally). The dorsalmost papilla in this ring lies at the center of the retina. In addition, up to seven small papillae lie within the ring. Bundles of fibers leave the neural retina via the papillae. These bundles remain separate as they pass through the distal portions of the neural retina and then merge before passing through the choroid. Bundles running through dorsal papillae receive fibers from a roughly wedge-shaped retinal area; bundles running through ventral papillae receive fibers from a small area of central retina and a disproportionately large area of peripheral retina. A band of high ganglion cell density was observed extending between the nasal and temporal poles of the retina. No correlation was found between the retinal areas contributing fibers to the bundles of axons emerging from individual papillae and the areas of high cell density. Furthermore, no correlation was found between the average area of the retinal ganglion cells and the ganglion cell density. From HRP preparations and axon counts we estimate that each retina of 95-mm catfish contains about 50,000 ganglion cells.
Assuntos
Peixes-Gato/anatomia & histologia , Ictaluridae/anatomia & histologia , Retina/anatomia & histologia , Animais , Disco Óptico/anatomia & histologia , Células Ganglionares da Retina/citologiaRESUMO
Horseradish peroxidase was applied to lesions in the optic nerve of catfish (Ictalurus punctatus). The retinae were processed to reveal HRP-labelled ganglion cells. The histochemical techniques employed allowed fine details of the dendritic arbor to be resolved. Flat-mounted retinae were examined and the following characteristics were noted in individual ganglion cells: Soma area, shape, and depth; number and diameter of major dendrites; shape, area, and depth(s) within the inner plexiform layer (ipl) of the dendritic arbor; origin of the axon (from the soma or a dendrite). On the basis of these characteristics, eleven classes of ganglion cells were delineated: four classes of giant cells (G1-G4) and seven classes of smaller cells (S1-S7). G1 cells had dendrites arborizing in the most distal sublamina of the ipl. G1 cells in the dorsal retina had nasotemporally elongated dendritic arbors. G2 cells had dendrites in the proximal portion of the ipl. G3 cells were almost completely confined to a band running between the nasal and temporal retinal poles, through the center of the retina. In this location, the cells had dorsoventrally elongated dendritic arbors, which were bistratified in the ipl. G4 cells were displaced into the inner nuclear layer. S1 and S4 cells had axons arising from their somata, and dendrites arborizing in the distal and the proximal ipl, respectively. S2 cells were typified by their unstratified dendritic arbors. Similarly, S3 cells were characterised by their bistratified arbors. S5 cells arborized in the most proximal ipl sublamina. S6 cells were small ganglion cells with their somata lying in the inner nuclear layer. S7 cells tended to have complex dendritic arbors, and their axons arose from dendrites.
Assuntos
Retina/citologia , Células Ganglionares da Retina/citologia , Vias Visuais/citologia , Animais , Dendritos/classificação , Peixes , Compressão Nervosa , Células Ganglionares da Retina/classificação , Degeneração RetrógradaRESUMO
Within the rat primary somatosensory cortex, neurons responding principally to movement of each individual mystacial vibrissa are grouped together in structures termed barrels. Previous studies have examined changes in the area of cortex showing increased 2-deoxyglucose uptake in response to vibrissal stimulation. These studies have shown that chronic removal of all but the central (C3) vibrissa in adult rats induces an enlarged representation of the remaining C3 barrel in the contralateral cortex. This increase is prevented by cortical norepinephrine depletion. The major question raised by such studies is whether such plasticity is due to structural rearrangement or unmasking of otherwise silent synapses. In this study, antibodies to GAP-43, a presynaptic protein whose synthesis is related to neuronal development and regeneration, were used to investigate this issue. In adult rat brain, tangential sections through layer IV of the barrel receptor field normally show moderate levels of GAP-43 immunoreactivity (GAP-IR) in the inter-barrel septa and low levels within the barrels themselves. The present study examined changes in the pattern of GAP-IR from 1 to 8 weeks after vibrissectomy with sparing of C3 as an index of possible physical reorganization of cortical circuits. Quantitative analysis of the cortices of animals with unilateral vibrissectomy with sparing of C3 showed that the area of low GAP-IR within the barrels surrounding C3 was decreased at 1 week (8.4% shrinkage; P less than 0.01) and 8 weeks (12.0% shrinkage; P less than 0.015), relative to the cortex ipsilateral to the surgery. Both bilateral vibrissectomy with sparing of C3 and ibotenic acid lesions of the ventrobasal thalamus produced similar results. Some evidence was also seen that the area of low GAP-IR in the C3 barrel shrank to a similar degree after such manipulations. Cortical norepinephrine depletion had no apparent effect on vibrissectomy-induced GAP-IR changes. These results suggest that removal of vibrissal input to the adult rat barrel cortex produces transynaptic induction of axonal sprouting within the barrel cortex.
Assuntos
Glicoproteínas de Membrana/análise , Regeneração Nervosa/fisiologia , Proteínas do Tecido Nervoso/análise , Plasticidade Neuronal , Ratos/fisiologia , Córtex Somatossensorial/química , Vibrissas/fisiologia , Vias Aferentes/fisiologia , Animais , Denervação , Proteína GAP-43 , Ácido Ibotênico/toxicidade , Locus Cerúleo/lesões , Locus Cerúleo/fisiologia , Masculino , Norepinefrina/deficiência , Norepinefrina/fisiologia , Ratos Endogâmicos , Córtex Somatossensorial/fisiologia , Tálamo/efeitos dos fármacosRESUMO
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.
Assuntos
Antagonistas de Receptores Adrenérgicos alfa 1 , Antagonistas Adrenérgicos alfa/farmacologia , Comportamento Animal/fisiologia , Lesões Encefálicas/psicologia , Inibidores da Captação Adrenérgica/farmacologia , Animais , Comportamento Animal/efeitos dos fármacos , Pressão Sanguínea/efeitos dos fármacos , Encéfalo/patologia , Lesões Encefálicas/patologia , Desipramina/farmacologia , Masculino , Norepinefrina/metabolismo , Norepinefrina/fisiologia , Prazosina/farmacologia , Ratos , Ratos Sprague-Dawley , Córtex Somatossensorial/metabolismo , Córtex Somatossensorial/patologiaRESUMO
Norepinephrine (NE), acting on brain adrenoceptors, plays an important role in barrel field neuronal activity and plasticity. For this reason, the distribution of alpha 1- and alpha 2-adrenoceptors in the somatosensory cortex barrel field was studied by autoradiographic techniques in rats undergoing plastic change or NE depletion. In layers IV and V of the cortex, the pattern of alpha 1-adrenoceptors (assessed by [3H]prazosin binding) varied across the barrel field. There was relatively low binding within the barrels themselves, with 21% higher binding in the surrounding septa. alpha 2-Adrenoceptor binding (assessed with [3H]paraminoclonidine) was almost homogeneous across the entire barrel field. Two weeks after noradrenergic deafferentation by unilateral lesioning of the locus coeruleus, there was a 16% upregulation of [3H]prazosin binding. This then returned to control levels of by 8 weeks. Peripheral deafferentation of sensory input to the barrel field produced the opposite effect on alpha 1-adrenoceptors. Unilateral removal of all but the central (C3) vibrissa (which induces plastic changes in the cortical representation of the spared virbrissa) caused a 12% decrease in [3H]prazosin binding in the whole barrel field at 2 weeks after surgery which returned to normal by 8 weeks. Therefore, alpha 1-adrenoceptors in the barrel field of the rat are affected in opposite ways by changes in NE content and afferent sensory input. We hypothesize that alpha 1-adrenoceptor levels are modulated after vibrissectomy through either an indirect reaction to reduced cortical gamma-aminobutyric acid levels, or by a reordering of metabolic priorities during plastic change of the cortical neuronal network.
Assuntos
Vias Aferentes/fisiologia , Neurônios/metabolismo , Norepinefrina/fisiologia , Receptores Adrenérgicos alfa 1/metabolismo , Córtex Somatossensorial/metabolismo , Vibrissas/inervação , Análise de Variância , Animais , Autorradiografia , Lateralidade Funcional , Locus Cerúleo/fisiologia , Masculino , Prazosina/metabolismo , Ratos , Ratos Sprague-Dawley , Receptores Adrenérgicos alfa 2/metabolismo , Fatores de Tempo , TrítioRESUMO
When the axons of goldfish retinal ganglion cells are severed the cell bodies undergo a series of changes as the axons regenerate. These changes begin to reverse when the axons start to innervate the tectum and by 3 months after the lesion the cell bodies have nearly returned to normal. When the axons projecting to the caudal tectum were severed by a mediolateral transection of the tectum, only retinal ganglion cells in the nasal portion of the contralateral retina underwent the changes normally associated with regeneration, followed by a speedy return to normal. Because the injured fibers probably did not fully retract from the tectum, these results indicated that: (1) the complete removal of the axons from the tectal milieu was not essential for initiating the cell body changes, and (2) close proximity to the target sites would speed the recovery of the cells. When the caudal portion of the tectum was ablated the retinal ganglion cells of the nasal retina remained enlarged significantly longer than after tectal transection. During the time the cells remained enlarged the electrophysiological projection onto the remaining rostral part of the tectum revealed no significant 'compression' of the visual field. Compression of the visual field onto the rostral portion of the tectum can be accelerated if the caudal tectal ablation is accompanied by an optic nerve crush. However, under this condition the recovery of ganglion cells in the nasal retina was significantly slower than the recovery of cells in the temporal retina. This may reflect an element of topographical specificity in the regulation of the recovery of the cell body from axonal injury.
Assuntos
Regeneração Nervosa , Nervo Óptico/fisiologia , Retina/fisiologia , Células Ganglionares da Retina/fisiologia , Colículos Superiores/fisiologia , Sinapses/fisiologia , Animais , Encéfalo/citologia , Encéfalo/fisiologia , Condutividade Elétrica , Carpa Dourada , Fatores de TempoRESUMO
Previous studies have shown that rats recover function after unilateral somatosensory cortex lesions, possibly by transfer of information processing to other brain areas not normally involved in those functions. In the present study, adult rats underwent unilateral contusions of the somatosensory cortex with ablation of the barrel receptor field. Behavioral testing with modified beam-walking and sensory neglect tasks demonstrated persistent somatosensory deficits in rats with left contusions but no apparent deficits in right injured animals. After 2 months, the [14C]2-deoxyglucose (2-DG) method was used to show the metabolic activity produced by unilateral stimulation of the facial vibrissae. In left injured animals, cortical metabolic activity rostral and caudal to the injury site was depressed both under basal conditions and during right vibrissal stimulation. On the other hand, comparison of the pattern of [14C]2-DG uptake in the intact, right cortex revealed changes in the pattern of glucose utilization associated with left injury combined with right vibrissal stimulation. Pattern changes were quantified by measuring the area in which glucose utilization was within the highest 25% of this range (high activity area; HAA). Right vibrissal stimulation in left injured rats caused an expansion of this HAA in the intact occipital/temporal cortex. Also, in the intact somatosensory cortex of left injured rats, there was an enlarged HAA whether or not vibrissal stimulation was performed. Thus, a combination of depressed peri-injury metabolic activity and aberrant activity in remote brain areas occurs following unilateral somatosensory cortex injury. It remains to be shown whether these factors ameliorate or contribute to persistent behavioral deficits.
Assuntos
Comportamento Animal/fisiologia , Concussão Encefálica/patologia , Concussão Encefálica/psicologia , Lateralidade Funcional/fisiologia , Córtex Somatossensorial/patologia , Animais , Autorradiografia , Química Encefálica/fisiologia , Desoxiglucose/metabolismo , Glucose/metabolismo , Masculino , Ratos , Ratos Sprague-Dawley , Vibrissas/fisiologiaRESUMO
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.
Assuntos
Transportadores de Cassetes de Ligação de ATP , Química Encefálica/efeitos dos fármacos , Glucose/metabolismo , Glibureto/metabolismo , Hipoglicemiantes/metabolismo , Obesidade/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização , Canais de Potássio/metabolismo , Receptores de Droga/metabolismo , Animais , Química Encefálica/fisiologia , Artérias Carótidas , Diuréticos Osmóticos/farmacologia , Ingestão de Energia , Glucose/farmacologia , Glibureto/farmacologia , Hipoglicemiantes/farmacologia , Injeções Intra-Arteriais , Masculino , Manitol/farmacologia , Canais de Potássio/fisiologia , Ratos , Ratos Sprague-Dawley , Receptores de Sulfonilureias , TrítioRESUMO
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.
Assuntos
Astrócitos/química , Axônios/química , Lesões Encefálicas/metabolismo , Neurônios/química , Animais , Astrócitos/citologia , Benzenossulfonatos , Benzoxazinas , Biomarcadores , Corantes , Proteína Glial Fibrilar Ácida/análise , Proteínas de Choque Térmico HSP72 , Proteínas de Choque Térmico/análise , Imuno-Histoquímica , Masculino , Degeneração Neural/fisiologia , Proteínas do Tecido Nervoso/análise , Proteínas de Neurofilamentos/análise , Neurônios/citologia , Neurônios/ultraestrutura , Oxazinas , Proteínas Proto-Oncogênicas c-fos/análise , Ratos , Ratos Sprague-Dawley , Fatores de TempoRESUMO
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.
Assuntos
Trifosfato de Adenosina/farmacologia , Encéfalo/efeitos dos fármacos , Hipotálamo/efeitos dos fármacos , Neurônios/química , Canais de Potássio/efeitos dos fármacos , Animais , Encéfalo/metabolismo , Catecolaminas/metabolismo , Hipotálamo/metabolismo , Masculino , Neuropeptídeo Y/metabolismo , Fenótipo , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Ácido gama-Aminobutírico/metabolismoRESUMO
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.
Assuntos
Glucose/farmacologia , Hipotálamo/metabolismo , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Obesidade/metabolismo , Animais , Núcleo Arqueado do Hipotálamo/citologia , Núcleo Arqueado do Hipotálamo/metabolismo , Dieta , Núcleo Hipotalâmico Dorsomedial/citologia , Núcleo Hipotalâmico Dorsomedial/metabolismo , Hipotálamo/citologia , Neurônios/química , Núcleo Hipotalâmico Paraventricular/citologia , Núcleo Hipotalâmico Paraventricular/metabolismo , Proteínas Proto-Oncogênicas c-fos/análise , Ratos , Ratos Mutantes , Ratos Sprague-Dawley , Núcleo Supraquiasmático/citologia , Núcleo Supraquiasmático/metabolismo , Núcleo Supraóptico/citologia , Núcleo Supraóptico/metabolismo , Núcleo Hipotalâmico Ventromedial/citologia , Núcleo Hipotalâmico Ventromedial/metabolismoRESUMO
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.
Assuntos
Concussão Encefálica/fisiopatologia , Edema Encefálico/fisiopatologia , Norepinefrina/metabolismo , Ácido 3,4-Di-Hidroxifenilacético/metabolismo , Antagonistas Adrenérgicos alfa/farmacologia , Animais , Química Encefálica/efeitos dos fármacos , Química Encefálica/fisiologia , Concussão Encefálica/metabolismo , Edema Encefálico/metabolismo , Tronco Encefálico/metabolismo , Tronco Encefálico/fisiopatologia , Cerebelo/metabolismo , Cerebelo/fisiopatologia , Hipotálamo/metabolismo , Hipotálamo/fisiopatologia , Indometacina/análogos & derivados , Indometacina/farmacologia , Masculino , Metoxi-Hidroxifenilglicol/metabolismo , Prazosina/farmacologia , Ratos , Ratos Sprague-Dawley , Córtex Somatossensorial/metabolismo , Córtex Somatossensorial/fisiopatologiaRESUMO
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.
Assuntos
Artérias Carótidas/fisiologia , Núcleo Hipotalâmico Dorsomedial/metabolismo , Glucose/administração & dosagem , Neurônios/metabolismo , Núcleo Hipotalâmico Paraventricular/metabolismo , Proteínas Proto-Oncogênicas c-fos/metabolismo , Núcleo Hipotalâmico Ventromedial/metabolismo , Animais , Núcleo Hipotalâmico Dorsomedial/citologia , Glucose/farmacologia , Imuno-Histoquímica , Injeções Intra-Arteriais , Masculino , Núcleo Hipotalâmico Paraventricular/citologia , Ratos , Ratos Sprague-Dawley , Núcleo Hipotalâmico Ventromedial/citologiaRESUMO
The brain contains neurons which alter their firing rates when ambient glucose concentrations change. An ATP-sensitive K+ (Katp) channel on these neurons closes and increases cell firing when ATP is produced by intracellular glucose metabolism. Binding of the antidiabetic sulfonylurea drugs to a site linked to this channel has a similar effect. Here rats with a propensity to develop diet-induced obesity (DIO) or to be diet-resistant (DR) when fed a diet moderately high in fat, energy and sucrose (HE diet) had low and high affinity sulfonylurea binding assessed autoradiographically with [3H]glyburide in the presence or absence of Gpp(NH)p. Before HE diet exposure, chow-fed DIO- and DR-prone rats were separated by their high vs. low 24 h urine NE levels. In DR-prone rats, low affinity [3H]glyburide binding sites comprised up to 45% of total binding with highest concentrations in the hypothalamus and amygdala. But DIO-prone rats had few or no low affinity binding sites throughout the forebrain. High affinity [3H]glyburide binding was similar between phenotypes. When rats developed DIO after 3 months on HE diet, their low affinity binding increased slightly. DR rats fed the HE diet gained the same amount of weight as chow-fed controls but their low affinity binding sites were reduced to DIO levels and both were significantly lower than chow-fed controls. By contrast, high affinity [3H]glyburide binding was increased in DR rats throughout the forebrain so that it significantly exceeded that in both DIO and chow-fed control rats. These studies demonstrate a significant population of low affinity sulfonylurea binding sites throughout the forebrain which, along with high affinity sites, are regulated as a function of both weight gain phenotype and diet composition.
Assuntos
Transportadores de Cassetes de Ligação de ATP , Encéfalo/efeitos dos fármacos , Dieta , Glucose/farmacologia , Obesidade/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização , Canais de Potássio/metabolismo , Receptores de Droga/metabolismo , Compostos de Sulfonilureia/metabolismo , Análise de Variância , Animais , Encéfalo/metabolismo , Suscetibilidade a Doenças , Masculino , Fenótipo , Ratos , Ratos Sprague-Dawley , Receptores de Sulfonilureias , Aumento de Peso/efeitos dos fármacosRESUMO
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.