RESUMO
Deer mice, Peromyscusmaniculatus, live at high altitudes where limited O2 represents a challenge to maintaining oxygen delivery to tissues. Previous work has demonstrated that hypoxia acclimation of deer mice and low altitude white-footed mice (P. leucopus) increases the force generating capacity of the diaphragm. The mechanism behind this improved contractile function is not known. Within myocytes, the myofilament plays a critical role in setting the rate and level of force production, and its ability to generate force can change in response to changes in physiological conditions. In the current study, we examined how chronic hypobaric hypoxia exposure of deer mice and white-footed mice influences the Ca2+ activation of force generation by skinned diaphragmatic myofilaments, and the phosphorylation of myofilament proteins. Results demonstrate that myofilament force production, and the Ca2+ sensitivity of force generation, were not impacted by acclimation to hypobaric hypoxia, and did not differ between preparations from the two species. The cooperativity of the force-pCa relationship, and the maximal rate of force generation were also the same in the preparations from both species, and not impacted by acclimation. Finally, the relative phosphorylation of TnT, and MLC was lower in deer mice than white-footed mice, but was not affected by acclimation. These results indicate that species differences in diaphragm function, and the increase in force production with hypoxia acclimation, are not due to differences, or changes, in myofilament function. However, it appears that diaphragmatic myofilament function in these species is not affected by chronic hypobaric hypoxia exposure.
Assuntos
Altitude , Diafragma/fisiologia , Hipóxia/metabolismo , Miofibrilas/fisiologia , Peromyscus/classificação , Peromyscus/fisiologia , Aclimatação , Animais , Cálcio/metabolismo , Especificidade da Espécie , Fatores de TempoRESUMO
AIM: We examined the effects of chronic hypoxia on diaphragm function in high- and low-altitude populations of Peromyscus mice. METHODS: Deer mice (P. maniculatus) native to high altitude and congeneric mice native to low altitude (P. leucopus) were born and raised in captivity to adulthood and were acclimated to normoxia or hypobaric hypoxia (12 or 9 kPa, simulating hypoxia at 4300 and 7000 m) for 6-8 weeks. We then measured indices of mitochondrial respiration capacity, force production, and fatigue resistance in the diaphragm. RESULTS: Mitochondrial respiratory capacities (assessed using permeabilized fibres with single or multiple inputs to the electron transport system), citrate synthase activity (a marker of mitochondrial volume), twitch force production, and muscle fatigue resistance increased after exposure to chronic hypoxia in both populations. These changes were not well explained by variation in the fibre-type composition of the muscle. However, there were several differences in diaphragm function in high-altitude mice compared to low-altitude mice. Exposure to a deeper level of hypoxia (9 kPa vs 12 kPa) was needed to elicit increases in mitochondrial respiration rates in highlanders. Chronic hypoxia did not increase the emission of reactive oxygen species from permeabilized fibres in highlanders, in contrast to the pronounced increases that occurred in lowlanders. In general, the diaphragm of high-altitude mice had greater capillary length densities, produced less force in response to stimulation and had shorter relaxation times. The latter was associated with higher activity of sarcoplasmic reticulum Ca2+ -ATPase (SERCA) activity in the diaphragm of high-altitude mice. CONCLUSION: Overall, our work suggests that exposure to chronic hypoxia increases the capacities for mitochondrial respiration, force production and fatigue resistance of the diaphragm. However, many of these effects are opposed by evolved changes in diaphragm function in high-altitude natives, such that highlanders in chronic hypoxia maintain similar diaphragm function to lowlanders in sea level conditions.
Assuntos
Aclimatação , Altitude , Diafragma/fisiopatologia , Hipóxia/fisiopatologia , Contração Muscular , Animais , Doença Crônica , Diafragma/metabolismo , Modelos Animais de Doenças , Metabolismo Energético , Hipóxia/metabolismo , Mitocôndrias Musculares/metabolismo , Fadiga Muscular , Força Muscular , Peromyscus , Espécies Reativas de Oxigênio/metabolismo , Especificidade da EspécieRESUMO
Voltage-gated chloride channels have recently been implicated as being important for cell proliferation and invasive cell migration of primary brain tumors cells. In the present study we provide several lines of evidence that glioma Cl- currents are primarily mediated by ClC-2 and ClC-3, two genes that belong to the ClC superfamily. Transcripts for ClC-2 thru ClC-7 were detected in a human glioma cell line by PCR, whereas only ClC-2, ClC-3, and ClC-5 protein could be identified by Western blot. Prominent ClC-2, -3, and -5 channel expression was also detected in acute patient biopsies from low- and high-grade malignant gliomas. Immunogold electron microscopic studies as well as digital confocal imaging localized a portion of these ClC channels to the plasma membrane. Whole-cell patch-clamp recordings show the presence of two pharmacologically and biophysically distinct Cl- currents that could be specifically reduced by 48 hr exposure of cells to channel-specific antisense oligonucleotides. ClC-3 antisense selectively and significantly reduced the expression of outwardly rectifying current with pronounced voltage-dependent inactivation. Such currents were sensitive to DIDS (200-500 microm) and 5-nitro-2-(3-phenylpropylamino) benzoic acid (165 microm). ClC-2 antisense significantly reduced expression of inwardly rectifying currents, which were potentiated by hyperpolarizing prepulses and inhibited by Cd2+ (200-500 microm). Currents that were mediated by ClC-5 could not be demonstrated. We suggest that ClC-2 and ClC-3 channels are specifically upregulated in glioma membranes and endow glioma cells with an enhanced ability to transport Cl-. This may in turn facilitate rapid changes in cell size and shape as cells divide or invade through tortuous extracellular brain spaces.
Assuntos
Astrocitoma/metabolismo , Canais de Cloreto/biossíntese , Glioblastoma/metabolismo , Glioma/metabolismo , Anticorpos/farmacologia , Astrocitoma/patologia , Biópsia , Western Blotting , Canais de Cloro CLC-2 , Membrana Celular/metabolismo , Canais de Cloreto/antagonistas & inibidores , Canais de Cloreto/genética , Cloretos/metabolismo , Glioblastoma/patologia , Glioma/patologia , Humanos , Imuno-Histoquímica , Oligonucleotídeos Antissenso/farmacologia , Técnicas de Patch-Clamp , Reação em Cadeia da Polimerase , RNA Mensageiro/biossíntese , Células Tumorais Cultivadas , Regulação para CimaRESUMO
The major classes of glial cells, namely astrocytes, oligodendrocytes, and microglial cells were compared in parallel for their susceptibility to damage after combined hypoxia and hypoglycemia or hypoxia alone. The three glial cell types were isolated from neonatal rat brains, separated, and incubated in N2/CO2-gassed buffer-containing glucose or glucose substitutes, 2-deoxyglucose or mannitol (both nonmetabolizable sugars). The damage to the cells after 6 hours' exposure was determined at 0, 1, 3, 7 days based on release of lactate dehydrogenase and counting of ethidium bromide-stained dead cells, double-stained with cell-type specific markers. When 2-deoxyglucose replaced glucose during 6 hours of hypoxia, both oligodendrocytes and microglia rarely survived (18% and 12%, respectively). Astroglia initially increased the release of lactate dehydrogenase but maintained 98% to 99% viability. When mannitol, a radical scavenger and osmolarity stabilizer, replaced glucose during 6 hours of hypoxia, oligodendrocytes rarely survived (10%), astroglia survival remained at 99%, but microglia survival increased to 50%. After exposure to 6 and 42 hours, respectively, of hypoxic conditions alone, oligodendrocytes exhibited 10% survival whereas microglia and astroglia were only temporarily stressed and subsequently survived. In conclusion, oligodendrocytes, then microglia, are the most vulnerable glial cell types in response to hypoxia or hypoglycemia conditions, whereas astrocytes from the same preparations recover.
Assuntos
Glucose/metabolismo , Microglia/metabolismo , Microglia/patologia , Oligodendroglia/metabolismo , Oligodendroglia/patologia , Animais , Morte Celular , Hipóxia Celular , Sobrevivência Celular , Células Cultivadas , Ratos , Ratos WistarRESUMO
The authors investigated the time course of leukocyte infiltration compared with microglial activation in adult rat brain slices after permanent middle cerebral artery occlusion (MCAO). To distinguish peripheral leukocytes from microglia, the blood cells were prelabeled in vivo with Rhodamine 6G (Rhod6G) i.v. before induction of ischemia. At specific times after infarct, invading leukocytes, microglia, and endothelial cells were labeled in situ with isolectin (IL)B4-FITC (ILB4). Six hours after MCAO only a few of the ILB4+ cells were colabeled by Rhod6G. These cells expressed the voltage-gated inwardly and outwardly rectifying K+ currents characteristic of macrophages. The majority of the ILB4+ cells were Rhod6G- and expressed a lack of voltage-gated channels, recently described for ramified microglial cells in brain slices, or exhibited only an inward rectifier current, a unique marker for cultured (but unstimulated) microglia. Forty-eight hours after MCAO, all blood-borne and the majority of Rhod6G- cells expressed outward and inward currents indicating that the intrinsic microglial population exhibited physiologic features of stimulated, cultured microglia. The ILB4+/Rhod6G- intrinsic microglial population was more abundant in the border zone of the infarct and their morphology changed from radial to ameboid. Within this zone, the authors observed rapidly migrating cells and recorded this movement by time-lapse microscopy. The current findings indicate that microglial cells acquire physiologic features of leukocytes at a later time point after MCAO.
Assuntos
Córtex Cerebral/irrigação sanguínea , Córtex Cerebral/citologia , Infarto da Artéria Cerebral Média/fisiopatologia , Leucócitos/citologia , Microglia/fisiologia , Animais , Isquemia Encefálica/imunologia , Isquemia Encefálica/metabolismo , Isquemia Encefálica/fisiopatologia , Movimento Celular/imunologia , Córtex Cerebral/fisiologia , Imunofenotipagem , Infarto da Artéria Cerebral Média/imunologia , Infarto da Artéria Cerebral Média/metabolismo , Leucócitos/química , Leucócitos/imunologia , Masculino , Potenciais da Membrana/imunologia , Microglia/química , Microglia/citologia , Microscopia de Vídeo , Técnicas de Cultura de Órgãos , Técnicas de Patch-Clamp , Potássio/fisiologia , Canais de Potássio/análise , Canais de Potássio/metabolismo , Ratos , Ratos WistarRESUMO
An indirect sandwich ELISA is described for the detection of bovine enteroviruses. The assay was developed as an alternative to the complement fixation test and proved to be more sensitive and convenient. Ten bovine enterovirus prototype strains were easily discriminated. No cross-reactions were observed with other picornaviruses including foot-and-mouth disease viruses, swine vesicular disease virus, porcine enteroviruses and bovine rhinovirus.
Assuntos
Enterovirus/isolamento & purificação , Ensaio de Imunoadsorção Enzimática/métodos , Animais , BovinosRESUMO
Fetal rat dorsal root ganglion neurons (7-8 days in culture) were labeled with [3H]arachidonic acid for 24 h. Stimulation with 10 microM bradykinin (BK) for 30 s resulted in nearly 2-fold increases in levels of radioactive diglyceride and arachidonic acid. A similar result was obtained in the absence of receptor stimulation using the Ca2+ channel agonist BAY K 8644 (10 microM, in the presence of 100 mM potassium chloride) or the Ca2+ ionophore, ionomycin (2.5 microM). If Ca2+ influx was inhibited by adding 3 mM Co2+, a blocker of voltage-sensitive calcium channels, or 2.5 mM EDTA, then BK-stimulated accumulation of both arachidonate and diglyceride was inhibited. These data suggest Ca2+ influx is required for ligand-stimulated accumulation of both arachidonate (a product of diglyceride-lipase or phospholipase A2) and diglyceride (a product of phospholipase C). Two distinct populations of channels may be involved in these reactions since pretreatment with 10 microM nifedipine or 50 microM verapamil (agents which block a subset of voltage-sensitive Ca2+ channels) inhibited BK-stimulated accumulation of arachidonic acid, but did not inhibit diglyceride accumulation. Such functional discrimination appears to have physiological importance; the inhibitory effect of nifedipine and verapamil on BK-stimulated arachidonate release was mimicked by pretreatment with peptides which decrease Ca2+ channel conductance in dorsal root ganglion neurons. The three peptides used were 1 microM neuropeptide Y, 10 microM somatostatin, and 10 microM [N-MePhe3,D-Pro4]-morphiceptin. The effect of neuropeptide Y was blocked by pretreatment with pertussis toxin.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Bradicinina/farmacologia , Endorfinas/farmacologia , Gânglios Espinais/fisiologia , Neurônios/fisiologia , Neuropeptídeo Y/farmacologia , Sistemas do Segundo Mensageiro/efeitos dos fármacos , Somatostatina/farmacologia , Analgésicos/farmacologia , Animais , Ácido Araquidônico , Ácidos Araquidônicos/metabolismo , Cálcio/fisiologia , Células Cultivadas , Diglicerídeos/metabolismo , Feto , Cinética , Modelos Biológicos , Neurônios/efeitos dos fármacos , Nifedipino/farmacologia , Toxina Pertussis , Ratos , Ratos Endogâmicos , Verapamil/farmacologia , Fatores de Virulência de Bordetella/farmacologiaRESUMO
This study details the novel application of predacious copepods, genus Mesocyclops, for control of Ochlerotatus tremulus (Theobald) group and Aedes aegypti (L.) mosquito larvae in subterranean habitats in north Queensland, Australia. During June 1997, 50 Mesocyclops sp. 1 were inoculated into one service manhole in South Townsville. Wet season rainfall and flooding in both 1998 and 2000 was responsible for the dispersal of copepods via the underground pipe system to 29 of 35 manholes over an area of 1.33 km2. Significant reductions in Aedes and Ochlerotatus larvae ensued. In these habitats, Mesocyclops and Metacyclops were able to survive dry periods, when substrate moisture content ranged from 13.8 to 79.9%. At the semiarid inland towns of Hughenden and Richmond, cracking clay soil prevents drainage of water from shallow service pits where Oc. tremulus immatures numbered from 292-18,460 per pit. Introduction of Mesocyclops copepods into these sites during May 1999 resulted in 100% control of Oc. tremulus for 18 mo. One uninoculated pit subsequently became positive for Mesocyclops with resultant control of mosquito larvae.
Assuntos
Aedes , Crustáceos , Culicidae , Controle Biológico de Vetores/métodos , Comportamento Predatório , Animais , QueenslandRESUMO
At semiarid Charters Towers, north Queensland, Australia, the importance of Aedes aegypti (L.) in wells was assessed in relation to the colonization of surface habitats during the wet season. From April to July 1999, 10 wells (five positive for Ae. aegypti) were monitored to assess their status and larvae population numbers therein. All surface containers located within a 100 m radius of each well were removed, treated with s-methoprene or sealed to prevent the utilization of these containers by mosquitoes. These inner cores were surrounded by outer zones for a further 100 m in which surface containers were left untreated but all subterranean habitats were treated. Ovitraps were monitored monthly in the inner cores for 36 wk from August 1999 to April 2000 and differences in the proportions of ovitraps positive for Ae. aegypti and Ochlerotatus notoscriptus (Skuse) were analyzed by logistic regression. Analysis of the proportions of ovitraps positive for Ae. aegypti near positive wells indicated significantly greater colonization from November to March (the wet season), compared with those situated near Ae. aegypti negative wells. As Oc. notoscriptus were not produced from subterranean sites, comparisons of the proportions of ovitraps positive for Oc. notoscriptus in positive and negative inner cores provided an indication of the relative productivity of the uncontrolled surface containers in the outer zones. Differences in the utililization of ovitraps by Oc. notoscriptus among positive and negative cores were observed during only one month (March), when oviposition was greater in ovitraps in the negative cores, compared with the positive cores. Best subsets linear regression analysis of the proportion of ovitraps positive forAe. aegypti against meteorological variables (rainfall, mean wind speed, mean relative humidity, mean minimum, and maximum temperature) during the week of ovitrapping indicated that minimum temperature and wind speed accounted for 63.4% of the variability. This study confirms that for semiarid towns such as Charters Towers, the practice of treating a relatively small number of key subterranean habitats during winter will significantly affect Ae. aegypti recolonization of surface container habitats during summer, the period of greatest risk for dengue.
Assuntos
Aedes/fisiologia , Animais , Larva , Oviposição , Estações do AnoRESUMO
In north Queensland, 14 localities were surveyed for mosquito larvae (third and fourth instar) during summer/autumn and winter from 1996 to 1999. Absolute population numbers in subterranean habitats, mainly service manholes and pits (97%) but also some wells, septic tanks, storm drains, and sumps, were expressed as a proportion of total numbers in these sites plus surface sites within a 100-m radius. When correction factors were applied to subterranean samples, the 472,477 larvae mainly of Aedes tremulus (Theobald) group, Aedes notoscriptus (Skuse), and Aedes aegypti (L.) comprised 78% of the total population. In relation to the proportion of the overall immature mosquito population from subterranean habitats (propsub), linear regression coefficients for minimum temperature, relative humidity, and Mesocyclops copepod prevalence were significant for winter data; but for summer, only relative humidity was significant. Linear regression coefficients for Mesocyclops prevalence approached significance (P = 0.061) in summer. When multiple linear regression was used to model propsub, 68% of the variation was accounted for by relative humidity and the prevalence of Mesocyclops. In the drier and cooler towns, increased use of subterranean sites during winter was caused by reduced availability of surface oviposition sites because of the dry season. In the wetter coastal towns, no such restrictions applied and ambient conditions remained more equitable all year round. Mesocyclops were surprisingly common, particularly in these coastal towns. Release of known numbers of Mesocyclops indicated that 3-sweep netting in service manholes was sensitive down to densities of one Mesocyclops per 10 liters, and overall recovery varied from 1 to 4%. In relation to control, service manholes represent a stable habitat for mosquito (7% positive overall) and Mesocyclops populations. If they remained wet, service manholes positive for mosquito immatures or Mesocyclops during summer/autumn had 96% and 85% chance, respectively, of being positive the following winter. Even allowing for the effect of drying, a mosquito-positive manhole had a 79% chance of remaining positive the following winter. In view of the importance of these sites as refuges from adverse ambient conditions, it is proposed that a winter control strategy using Mesocyclops presents a cost-effective control option to reduce the recolonization of surface sites when conditions become more suitable.
Assuntos
Aedes/fisiologia , Arbovírus , Culex/fisiologia , Insetos Vetores/fisiologia , Controle de Mosquitos , Animais , Calibragem , Meio Ambiente , Queensland , Sensibilidade e EspecificidadeRESUMO
Virgin female Microtus ochrogaster were exposed to paired and unpaired unfamiliar adult males in semi-natural arenas. Although females were reproductively activated by both types of males, more than twice as many were activated by unpaired (51.6%) as by paired (18.2%) males. Our results suggest that, in natural populations of prairie voles, the proportion of philopatric females becoming reproductive is related to the abundance of unpaired males within the population.
RESUMO
BACKGROUND: A 44-year-old woman came to us with a chief symptom of "jumping letters side-to-side, which is most noticeable while reading." The onset occurred after she had experienced a closed head traumatic brain injury 3 years earlier. Several neuro-ophthalmologists diagnosed a fixational instability secondary to saccadic intrusions and prescribed Gabapentin, which provided minimal relief. METHODS: The term saccadic intrusions refers to an inappropriate saccade with a disrupting effect on fixation. Our examination revealed a myopic anisometropia. Motility testing confirmed saccadic intrusions that lessened on occlusion of either eye and superior gaze. A plano spectacle with six-prism diopter-yoked base down was used to position the eyes in the superior null point. Electro-oculography, using the Visagraph II, demonstrated pre and post changes with the prism. The uncorrected anisometropia allowed the patient to be monocular under binocular viewing conditions. CASE REPORT: The case report focuses on fixational problems that may occur secondary to traumatic brain injuries. There is evidence that the origin of the problem may be from uninhibited brain stem circuits. Pharmacological treatment may only offer transient improvement. The responsibility of a functional cure is often placed on the optometrist. CONCLUSION: This case demonstrates how an alternative use of prism and prescription application can play an important role in the management of fixation dysfunctions.
Assuntos
Óculos , Transtornos da Motilidade Ocular/terapia , Movimentos Sacádicos , Acidentes de Trânsito , Adulto , Lesões Encefálicas/complicações , Feminino , Humanos , Transtornos da Motilidade Ocular/etiologia , Transtornos da Motilidade Ocular/fisiopatologia , Acuidade VisualRESUMO
Paraquat, bipyridilium herbicide know to cause pulmonary fibrosis, was injected IP into rats. The rate of synthesis of collagen by lung minces from these and control rats was evaluated by measuring the rate of synthesis of hydroxyproline, a specific marker for collagen in lung. Synthesis was measured by incubating lung minces with radioactive proline for various amounts of time, after which proline specific activity and labeled hydroxyproline were determined. The size of the proline pool within the lung minces was significantly elevated in minces from rats that had been injected with paraquat, thus causing the specific activity of the [3H]proline precursor to be lower in these lungs than in those from control animals. Lung minces from rats administered paraquat made more collagen than did those from uninjected controls. The actual increase in rate of collagen synthesis correlated well with other independent estimates of paraquat-induced damage to the lungs.
Assuntos
Colágeno/biossíntese , Pulmão/efeitos dos fármacos , Paraquat/farmacologia , Animais , Hidroxiprolina/biossíntese , Técnicas In Vitro , Pulmão/metabolismo , Prolina/metabolismo , Fibrose Pulmonar/induzido quimicamente , RatosRESUMO
1. The sorbitol dehydrogenases [L-iditol: NAD oxidoreductase] from livers of cow, man, rat and sheep each possess molecular weights of about 140,000. The beef, rat and sheep liver enzymes are composed of subunits of molecular weight 40,000. 2. The sorbitol dehydrogenases from livers of these four species each possess an isoelectric point of 7.3. 3. The four enzyme preparations show identical mobilities upon disc-gel electrophoresis and yield a single band of enzymic activity. 4. Sorbitol dehydrogenase activity is activated by the presence of ampholines or by increasing ionic strengths, with maximal activation at about 0.5 M salt concentration. These factors may cause the Km for NAD to be lowered.
Assuntos
L-Iditol 2-Desidrogenase/isolamento & purificação , Fígado/enzimologia , Mamíferos/metabolismo , Desidrogenase do Álcool de Açúcar/isolamento & purificação , Animais , Bovinos , Humanos , Ponto Isoelétrico , Cinética , L-Iditol 2-Desidrogenase/metabolismo , Peso Molecular , Conformação Proteica , Ratos , Ovinos , Especificidade da EspécieRESUMO
Schwann cells establish close contact with axons during development, and this is maintained throughout life. Signaling by neurotransmitters may play an important role in Schwann cell-axon interaction. Schwann cells were examined for the presence of neuroligland receptors that are linked to increases in levels of cytoplasmic calcium. Schwann cell cultures were prepared from neonatal rat sciatic nerve and, after 0.25, 1, 4, 7, and 14 days in vitro (DIV), loaded with the calcium indicator dye fura 2-AM. The influence of neuroligands on the cytosolic free calcium concentration ([Ca2+]i) was then examined at each time point using a video-based imaging system. Approximately 80-95% of all freshly isolated Schwann cells responded to 10 microM ATP with a three-fold rise in [Ca2+]i. Bradykinin, glutamate, and histamine had no or only partial and inconsistent responses. The ATP-induced calcium response disappeared within 4 DIV. Culturing cells in the presence of cyclic AMP (cAMP) analogues (which induce proliferation and differentiation in vitro) restored the ability of Schwann cells to respond to ATP with increased [Ca2+]i. In the presence of cAMP analogues the extent of recovery of ATP responsiveness was dependent on serum concentration. Fifty to ninety percent of cells regained calcium responsiveness to ATP when grown in medium containing cAMP analogues and 1% serum. These cells also exhibited immunoreactivity to P0 antibody, characteristic of the myelinating lineage. In contrast, only 15-30% of the Schwann cells regained calcium responsiveness when grown in medium containing cAMP analogues and 10% serum.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Cálcio/metabolismo , AMP Cíclico/análogos & derivados , AMP Cíclico/farmacologia , Receptores Purinérgicos P2/fisiologia , Células de Schwann/metabolismo , Nervo Isquiático/metabolismo , Trifosfato de Adenosina/farmacologia , Animais , Animais Recém-Nascidos , Bradicinina/farmacologia , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Fura-2 , Histamina/farmacologia , Cinética , Ratos , Receptores Purinérgicos P2/efeitos dos fármacos , Células de Schwann/citologia , Células de Schwann/efeitos dos fármacos , Espectrometria de Fluorescência , Fatores de Tempo , Regulação para Cima/efeitos dos fármacosRESUMO
We addressed the question of whether glial cells in intact white matter tracts express neurotransmitter receptors and we used Ca+2 signalling as a probe to detect the receptor activation. Corpus callosum slices from postnatal mice were bulk-loaded with the Ca+2-sensitive fluorescent dye fluo-3, and confocal microscopy was used to measure Ca+2 transients in response to neuroligands. Glial cell bodies were intensely dye-loaded and could be discriminated from the diffuse fluorescence of axons. Subpopulations of glial cells from slices obtained at postnatal days 3 to 7 responded with Ca+2 signals to ATP, glutamate, histamine, GABA, norepinephrine, serotonin, angiotensin II, bradykinin, and substance P. These subpopulations showed a distinct overlap; cells which were responsive to substance P always showed Ca+2 signalling in response to histamine, ATP, GABA and high K+ (membrane depolarization). GABA-responsive cells almost always showed a [Ca+2]i increase after membrane depolarization. In brain slices from postnatal day 11 to 18 animals, the Ca+2 responses were evident for glutamate, ATP, and norepinephrine, while GABA, substance P, serotonin, histamine, or angiotensin II rarely elicited a response. This study demonstrates that white matter glial cells in slices exhibit a large repertoire of neurotransmitter responses linked to Ca+2 signalling and that these receptor systems are differentially distributed on sub-populations of glial cells.
Assuntos
Cálcio/fisiologia , Corpo Caloso/fisiologia , Neuroglia/fisiologia , Transdução de Sinais/fisiologia , Animais , Astrócitos/metabolismo , Linhagem da Célula , Corpo Caloso/citologia , Eletrofisiologia , Fluorometria , Potenciais da Membrana/fisiologia , Camundongos , Oligodendroglia/metabolismo , Técnicas de Patch-Clamp , Receptores de Aminoácido/biossíntese , Receptores de Neuropeptídeos/biossíntese , Receptores de Neurotransmissores/biossíntese , Receptores Purinérgicos P1/biossíntese , Receptores Purinérgicos P2/biossínteseRESUMO
Schwann cells freshly isolated from the sciatic nerves of neonatal rats respond to exogenously applied ATP with a rapid increase in cytosolic calcium. This increase in [Ca2+]i is mediated by a P2Y-purinergic pathway (Lyons et al.: J. Neurochem. 63:552-560, 1994) and was measured using the calcium indicator dye, fura-2/AM, and a video-enhanced calcium imaging system. The ability to respond to ATP with increases in intracellular calcium is lost over a period of several days in culture; this loss can be prevented or reversed by application of cAMP analogs in a defined medium. We now demonstrate that the direct contact of Schwann cells with neurons also induces and stabilizes this ATP responsiveness. The induction of ATP responsiveness was observed among all Schwann cells contacting neurites, including those forming myelin, and regardless of whether the source of neurons was dorsal root ganglion neurons or superior cervical ganglion neurons. Approximately 85% of Schwann cells responded to ATP over the time studied (72 d in coculture). Addition of axolemma to Schwann cell cultures did not induce ATP responsiveness. We also examined the ATP responsiveness of Schwann cells in situ (excised nerves) using laser-scanning confocal microscopy and the calcium indicator dye, fluo-3/AM. Schwann cells in intact sciatic nerve segments isolated from neonatal and 16-day-old rats exhibited ATP-mediated [Ca2+]i increases. We conclude that neuronal contact is necessary for the expression of the ATP-mediated calcium responses in Schwann cells and that these responses are independent of myelin formation or maintenance.
Assuntos
Trifosfato de Adenosina/farmacologia , Cálcio/metabolismo , Comunicação Celular , Neurônios/fisiologia , Células de Schwann/metabolismo , Animais , Animais Recém-Nascidos , Axônios/fisiologia , Transporte Biológico Ativo/efeitos dos fármacos , Canais de Cálcio/metabolismo , Membrana Celular/fisiologia , Células Cultivadas , Gânglios Espinais/citologia , Gânglios Espinais/embriologia , Microscopia Confocal , Bainha de Mielina/fisiologia , Ratos , Ratos Sprague-Dawley , Células de Schwann/efeitos dos fármacos , Nervo Isquiático/citologia , Gânglio Cervical Superior/citologiaRESUMO
BACKGROUND: Hyperopia is the most common refractive error of children. Children with mild (or even moderate) levels of hyperopia usually do not experience visual problems resulting from this hyperopia. However, children with moderate-to-high degrees of hyperopia are at significantly increased risk for the development of amblyopia and strabismus. It is this association with these visually threatening disorders that makes hyperopia in children an important public health problem. In addition, even lesser degrees of hyperopia may affect the child's ability to perform well in near-related tasks, such as reading. The effect hyperopia has on an individual child is dependent on a variety of factors, including the magnitude of hyperopia, the age of the individual, the status of the accommodative and convergence system, and the demands placed on the visual system. Early detection and treatment of hyperopia may help prevention of potential complications from adversely impacting the child's vision. Although much is known about childhood hyperopia and its effects on vision, there is also much that is not known. The natural history, ocular biometry, relationship to accommodative function, the indications for treatment, and the most effective treatment modalities are among the underlying issues and clinical considerations awaiting more complete understanding.
Assuntos
Hiperopia , Acomodação Ocular , Ambliopia/etiologia , Ambliopia/prevenção & controle , Criança , Pré-Escolar , Humanos , Hiperopia/complicações , Hiperopia/diagnóstico , Hiperopia/terapia , Lactente , Optometria/métodos , Refração Ocular , Estrabismo/etiologia , Estrabismo/prevenção & controleRESUMO
1. The acidic and neutral glycoproteins secreted by cultured tracheal explants from pigs, sheep, rats, mice, monkeys, guinea pigs, dogs and chickens were purified and fractionated by column chromatography on DEAE-cellulose and by electrophoresis on cellulose acetate. 2. The ratios of acidic to neutral mucus glycoproteins were compared for the above animals with that of mucus glycoproteins secreted by cultured human bronchi. 3. The observed ratios of acidic to neutral glycoproteins ranged from 4.0 (mouse) to 7.2 (chicken and pig) from cultured tracheae; secreted human bronchial mucus had a ratio of 2.7. 4. The ratio of acidic to neutral glycoproteins secreted by tracheal explants varied with duration of incubation of the trachea in culture.
Assuntos
Glicoproteínas/metabolismo , Traqueia/metabolismo , Animais , Brônquios/metabolismo , Galinhas , Cromatografia DEAE-Celulose , Cães , Eletroforese em Acetato de Celulose , Glicoproteínas/análise , Cobaias , Humanos , Concentração de Íons de Hidrogênio , Macaca mulatta , Camundongos , Muco/metabolismo , Ratos , Ovinos , SuínosRESUMO
Neocortical freeze lesions have been widely used to study neuronal mechanisms underlying hyperexcitability in dysplastic cortex. Comparatively little attention has been given to biophysical changes in the surrounding astrocytes that show profound morphological and biochemical alterations, often referred to as reactive gliosis. Astrocytes are thought to aid normal neuronal function by buffering extracellular K(+). Compromised astrocytic K(+) buffering has been proposed to contribute to neuronal dysfunction. Astrocytic K(+) buffering is mediated, partially, by the activity of inwardly rectifying K(+) channels (K(IR)) and may involve intracellular redistribution of K(+) through gap-junctions. We characterized K(+) channel expression and gap-junction coupling between astrocytes in freeze-lesion-induced dysplastic neocortex. Whole cell patch-clamp recordings were obtained from astrocytes in slices from postnatal day (P) 16--P24 rats that had received a freeze-lesion on P1. A marked increase in glial fibrillary acidic protein immunoreactivity was observed along the entire length of the freeze lesion. Clusters of proliferative (bromo-deoxyuridine nuclear staining, BrdU+) astrocytes were seen near the depth of the microsulcus. Astrocytes in cortical layer I surrounding the lesion were characterized by a significant reduction in K(IR). BrdU-positive astrocytes near the depth of the microsulcus showed essentially no expression of K(IR) channels but markedly enhanced expression of delayed rectifier K(+) (K(DR)) channels. These proliferative cells showed virtually no dye coupling, whereas astrocytes in the hyperexcitable zone adjacent to the microsulcus displayed prominent dye-coupling as well as large K(IR) and outward K(+) currents. These findings suggest that reactive gliosis is accompanied by a loss of K(IR) currents and reduced gap junction coupling, which in turn suggests a compromised K(+) buffering capacity.