RESUMO
BACKGROUND AND STUDY AIMS: Hypoxic hepatitis (HH) is an acute liver injury that develops in patients with underlying diseases, such as heart failure, respiratory failure, septic/toxic shock. However, some patients do not have underlying diseases or episodes which are known to result in HH. Here, we analyzed the clinical characteristics of this particular patient group (called 'unknown HH' hereafter) to understand its pathogenesis. PATIENTS AND METHODS: Between October 2010 and January 2016, 157 consecutive patients with acute liver injury were admitted to our hospital. Among these patients, 15 patients were categorized as unknown HH. Medical histories and blood test results of unknown HH were analyzed. RESULTS: Among 15 patients of unknown HH, 11 were habitual drinkers and all experienced one of digestive symptoms which might result in mild hypovolemia such as vomiting, diarrhea, appetite loss, and epigastralgia. All patients of unknown HH presented marked elevation of serum ferritin concentration paralleled with aspartate transaminase (AST), alanine transaminase (ALT), and lactate dehydrogenase (LDH) concentrations. The serum levels of ferritin, ALT, LDH, and prothrombin time-international normalized ratio (PT-INR) were rapidly decreased during hospitalization and all 15 patients of unknown HH recovered without any complication. CONCLUSIONS: We found the particular group of HH with marked elevation of serum ferritin probably due to intrahepatic macrophage activation. Anti-inflammatory treatments might be effective for this group of hypoxic hepatitis.
Assuntos
Hepatite , Alanina Transaminase , Aspartato Aminotransferases , Ferritinas , Humanos , MacrófagosRESUMO
We introduce our technique for the treatment of aneurysms arising in the descending thoracic aorta and the thoracoabdominal aorta. Thoracotomy is performed at a single site. The costal arch is transected to ensure an adequate field of vision. A lifting hook is used to open the proximal side of the aorta. The diaphragm is not totally transected to preserve respiratory function after surgery. In principle, partial extracorporeal circulation is performed using a percutaneous cardiopulmonary support system. The dose of heparin for systemic treatment is limited to 50 U/kg. The abdominal branches are perfused with the use of balloon catheters. Cardiac arrest is induced for about 10 seconds by intravenous administration of adenosine triphosphate to avoid aortic injury when the proximal aorta is clamped during partial extracorporeal circulation and to prevent massive bleeding when the elephant trunk is clamped. To prevent paraplegia, the Adamkiewics artery and 2 pairs of adjacent intercostal arteries identified by preoperative computed tomography are reconstructed, and cerebrospinal drainage and motor evoked potential monitoring are performed.
Assuntos
Aneurisma da Aorta Abdominal/cirurgia , Aneurisma da Aorta Torácica/cirurgia , Humanos , Procedimentos Cirúrgicos Vasculares/métodosRESUMO
Recently, several groups have developed green fluorescent protein (GFP)-based Ca(2+) probes. When applied in cells, however, these probes are difficult to use because of a low signal-to-noise ratio. Here we report the development of a high-affinity Ca(2+) probe composed of a single GFP (named G-CaMP). G-CaMP showed an apparent K(d) for Ca(2+) of 235 nM. Association kinetics of Ca(2+) binding were faster at higher Ca(2+) concentrations, with time constants decreasing from 230 ms at 0.2 microM Ca(2+) to 2.5 ms at 1 microM Ca(2+). Dissociation kinetics (tau approximately 200 ms) are independent of Ca(2+) concentrations. In HEK-293 cells and mouse myotubes expressing G-CaMP, large fluorescent changes were observed in response to application of drugs or electrical stimulations. G-CaMP will be a useful tool for visualizing intracellular Ca2+ in living cells. Mutational analysis, together with previous structural information, suggests the residues that may alter the fluorescence of GFP.
Assuntos
Cálcio/análise , Cálcio/metabolismo , Proteínas Luminescentes/metabolismo , Trifosfato de Adenosina/farmacologia , Sequência de Aminoácidos , Animais , Carbacol/farmacologia , Linhagem Celular , Galinhas , Ácido Edético/farmacologia , Proteínas de Fluorescência Verde , Humanos , Indicadores e Reagentes , Ionomicina/farmacologia , Rim , Cinética , Proteínas Luminescentes/análise , Proteínas Luminescentes/genética , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Quinase de Cadeia Leve de Miosina/genética , Quinase de Cadeia Leve de Miosina/metabolismo , Miosinas/química , Miosinas/genética , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/metabolismo , Proteínas Recombinantes/análise , Proteínas Recombinantes/metabolismo , TransfecçãoRESUMO
Seizure is a form of excessive neuronal excitation and seizure-induced neuronal damage has profound effects on the prognosis of epilepsy. In various seizure models, the inactivation of Ca2+/calmodulin-dependent protein kinase II (CaMKII) occurs during seizure activity preceding neuronal cell death. CaMKII is a multifunctional protein kinase enriched in the brain and involved in various ways the regulation of neuronal activity. CaMKII inactivation during seizure activity may modify neuronal cell survival after seizure. However, the mechanism for CaMKII inactivation and its consequence after seizure recovery remain to be elucidated yet. In the present study, we employed a prolonged seizure model by systemic injection of kainic acid into rats and biochemically examined the activity state of CaMKII. In status epilepticus induced by kainic acid, not only the inactivation of CaMKII in brain homogenate, but also a shift in the distribution of CaMKII protein from the soluble to particulate fraction occurred in both hippocampus and parietal cortex. The particulate CaMKII showed a large decrease in the specific activity and a concurrent large increase in the autophosphorylation ratio at Thr-286 (alpha) and at Thr-287 (beta). In contrast, the soluble CaMKII showed normal or rather decreased specific activity and autophosphorylation ratio. After 24 h of recovery from kainic acid-induced status epilepticus, all such changes had disappeared. On the other hand, the total amount of CaMKII was decreased by 35% in hippocampus and 20% in parietal cortex, but the existing CaMKII was indistinguishable from those of controls in terms of the autonomous activity ratio, specific activity and autophosphorylation ratio. Thus, CaMKII inactivation in kainic acid-induced status epilepticus seems to be derived not from simple degradation of the enzyme, but from the formation of the autophosphorylated, inactivated and sedimentable CaMKII. Such a form of CaMKII may be important during pathological conditions in vivo in preventing excessive CaMKII activation due to Ca2+ overload.
Assuntos
Encéfalo/enzimologia , Sinalização do Cálcio/fisiologia , Proteínas Quinases Dependentes de Cálcio-Calmodulina/metabolismo , Cálcio/metabolismo , Epilepsia/enzimologia , Recuperação de Função Fisiológica/fisiologia , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/fisiopatologia , Sinalização do Cálcio/efeitos dos fármacos , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Convulsivantes/farmacologia , Modelos Animais de Doenças , Ativação Enzimática/efeitos dos fármacos , Ativação Enzimática/fisiologia , Epilepsia/induzido quimicamente , Epilepsia/fisiopatologia , Ácido Caínico/farmacologia , Masculino , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Fosforilação/efeitos dos fármacos , Ratos , Ratos Wistar , Recuperação de Função Fisiológica/efeitos dos fármacos , Solubilidade , Estado Epiléptico/induzido quimicamente , Estado Epiléptico/enzimologia , Estado Epiléptico/fisiopatologia , Treonina/metabolismoRESUMO
During neuronal differentiation and maturation, electrical excitability is essential for proper gene expression and the formation of synapses. The expression of ion channels is crucial for this process; in particular, voltage-gated K(+) channels function as the key determinants of membrane excitability. Previously, we reported that the A-type K(+) current (I(A)) and Kv4.2 K(+) channel subunit expression increased in cultured cerebellar granule cells with time. To examine the correlation between ion currents and the action potential, in the present study, we measured developmental changes of action potentials in cultured granule cells using the whole-cell patch-clamp method. In addition to an observed increment of I(A), we found that the Na(+) current also increased during development. The increase in both currents was accompanied by a change in the membrane excitability from the nonspiking type to the repetitive firing type. Next, to elucidate whether Kv4.2 is responsible for the I(A) and to assess the effect of Kv4 subunits on action potential waveform, we transfected a cDNA encoding a dominant-negative mutant Kv4.2 (Kv4.2dn) into cultured cells. Expression of Kv4.2dn resulted in the elimination of I(A) in the granule cells. This result demonstrates that members of the Kv4 subfamily are responsible for the I(A) in developing granule cells. Moreover, elimination of I(A) resulted in shortening of latency before the first spike generation. In contrast, expression of wild-type Kv4.2 resulted in a delay in latency. This indicates that appearance of I(A) is critically required for suppression of the excitability of granule cells during their maturation.
Assuntos
Cerebelo/fisiologia , Neurônios/fisiologia , Canais de Potássio de Abertura Dependente da Tensão da Membrana , Canais de Potássio/fisiologia , Potenciais de Ação/fisiologia , Algoritmos , Animais , Linhagem Celular , Cerebelo/citologia , Grânulos Citoplasmáticos/fisiologia , Eletrofisiologia , Proteínas de Fluorescência Verde , Humanos , Cinética , Proteínas Luminescentes , Potenciais da Membrana/genética , Potenciais da Membrana/fisiologia , Camundongos , Mutação/genética , Mutação/fisiologia , Oligonucleotídeos/genética , Técnicas de Patch-Clamp , Canais de Potássio/genética , Canais de Potássio ShalRESUMO
Spinocerebellar ataxia 6 (SCA6) is caused by expansion of a polyglutamine stretch, encoded by a CAG trinucleotide repeat, in the human P/Q-type Ca(2+) channel alpha(1A) subunit. Although SCA6 shares common features with other neurodegenerative glutamine repeat disorders, the polyglutamine repeats in SCA6 are exceptionally small, ranging from 21 to 33. Because this size is too small to form insoluble aggregates that have been blamed for the cause of neurodegeneration, SCA6 is the disorder suitable for exploring the pathogenic mechanisms other than aggregate formation, whose universal role has been questioned. To characterize the pathogenic process of SCA6, we studied the effects of polyglutamine expansion on channel properties by analyzing currents flowing through the P/Q-type Ca(2+) channels with an expanded stretch of 24, 30, or 40 polyglutamines, recombinantly expressed in baby hamster kidney cells. Whereas the Ca(2+) channels with =24 polyglutamines showed normal properties, the Ca(2+) channels with 30 or 40 polyglutamines exhibited an 8 mV hyperpolarizing shift in the voltage dependence of inactivation, which considerably reduces the available channel population at a resting membrane potential. The results suggest that polyglutamine expansion in SCA6 leads to neuronal death and cerebellar atrophy through reduction in Ca(2+) influx into Purkinje cells and other neurons. Besides the widely accepted notion that polyglutamine stretches exert toxic effects by forming aggregates, expanded polyglutamines directly alter functions of the affected gene product.
Assuntos
Canais de Cálcio Tipo P/fisiologia , Canais de Cálcio Tipo Q/fisiologia , Peptídeos/química , Animais , Apoptose , Canais de Cálcio Tipo P/química , Canais de Cálcio Tipo P/metabolismo , Canais de Cálcio Tipo Q/química , Canais de Cálcio Tipo Q/metabolismo , Linhagem Celular , Cricetinae , Humanos , Potenciais da Membrana , Técnicas de Patch-Clamp , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Ataxias Espinocerebelares/metabolismoRESUMO
We investigated whether the specific binding or labeling of 125I-omega-CgTX on crude membranes from chick whole brain was affected when endogenous GTP binding protein (G protein) was activated by GTP analogues, mastoparan (MP) and aluminum fluoride (AIF4-; AICl3 + NaF). Both GTPgammaS and Gpp(NH)p attenuated the inhibitory effect of selective N-type Ca channel inhibitors such as aminoglycoside antibiotics (AGs) or dynorphine (1-13)(Dyn) on specific 125I-omega-CgTX binding in a dose-dependent manner. On the other hand, the inhibitory effects of the divalent metal cations Cd2+, Co2+, Mg2+ and Mn2- on such binding were not attenuated by GTPgammaS. MP and AIF4- also attenuated the inhibitory effect of Neo on this binding similar to GTPgammaS. The attenuating effect of MP was enhanced by the presence of Mg2+ in a dose-dependent manner. However, GTP analogues, MP and AIF4-, did not affect binding or labeling without AGs or Dyn. GTPgammaS, MP and AIF4- also attenuated the specific labeling of a 215-kDa band in crude membranes with 125I-omega-CgTX using the cross-linker DSS (non-reduced condition) in the presence of Neo. These results indicate that there are direct or indirect relationships between N-type Ca channels and G proteins via binding sites for AGs or MP.
Assuntos
Compostos de Alumínio/farmacologia , Fluoretos/farmacologia , Proteínas de Ligação ao GTP/metabolismo , Guanosina Trifosfato/análogos & derivados , Peptídeos/metabolismo , Venenos de Vespas/farmacologia , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Galinhas , Peptídeos e Proteínas de Sinalização Intercelular , Radioisótopos do Iodo , ômega-Conotoxina GVIARESUMO
The relation between spontaneous contraction, Ca2+ oscillations, and sarcoplasmic reticulum (SR) function was studied in cultured neonatal rat cardiac myocytes. Spontaneous contraction and Ca2+ oscillations were irregular at day 2 of culture but became regular at day 6 of culture in neonatal rat cardiac myocytes. The rate of spontaneous contraction and the frequency of Ca2+ oscillations were decreased by verapamil and were abolished in the absence of extracellular Ca2+ at both day 2 and day 6 of culture. Ryanodine and thapsigargin increased the rate of contraction and the frequency of Ca2+ oscillations at day 2 of culture but did not affect contractions and Ca2+ oscillations at day 6 of culture. Ultrastructural observation showed that the structure of SR developed less at day 6 of culture. The present results suggest that spontaneous contraction and Ca2+ oscillations are due mainly to extracellular Ca2+ influx but not to Ca2+ release from SR in neonatal rat cardiac myocytes.
Assuntos
Coração/fisiologia , Contração Miocárdica/fisiologia , Retículo Sarcoplasmático/fisiologia , Animais , Animais Recém-Nascidos , Cálcio/metabolismo , Cálcio/farmacologia , Células Cultivadas , Miocárdio/citologia , Miocárdio/metabolismo , Ratos , Ratos Wistar , Rianodina/farmacologia , Retículo Sarcoplasmático/metabolismo , Tapsigargina/farmacologiaRESUMO
We have developed a novel method that uses a microfilter mask to produce ultraviolet-induced DNA lesions in localized areas of the cell nucleus. This technique allows us to visualize localized DNA repair in situ using immunologic probes. Two major types of DNA photoproducts [cyclobutane pyrimidine dimers and (6-4) photoproducts] were indeed detected in several foci per nucleus in normal human fibroblasts. They were repaired at those localized sites at different speeds, indicating that DNA photoproducts remain in relatively fixed nuclear positions during repair. A nucleotide excision repair protein, proliferating cell nuclear antigen, was recruited to the sites of DNA damage within 30 min after ultraviolet exposure. The level of proliferating cell nuclear antigen varied with DNA repair activity and diminished within 24 h. In contrast, almost no proliferating cell nuclear antigen fluorescence was observed within 3 h in xeroderma pigmentosum fibroblasts, which could not repair either type of photolesion. These results demonstrate that this technique is useful for visualizing the normal nucleotide excision repair process in vivo. Interestingly, however, in xeroderma pigmentosum cells, proliferating cell nuclear antigen appeared at ultraviolet damage sites after a delay and persisted as late as 72 h after ultraviolet exposure. This result suggests that this technique is also valuable for examining an incomplete or stalled nucleotide excision repair process caused by the lack of a single functional nucleotide excision repair protein. Thus, the technique provides a powerful approach to understanding the temporal and spatial interactions between DNA damage and damage-binding proteins in vivo.
Assuntos
Dano ao DNA/efeitos da radiação , Reparo do DNA , Fibroblastos/fisiologia , Fibroblastos/efeitos da radiação , Raios Ultravioleta , Linhagem Celular , Núcleo Celular/fisiologia , Núcleo Celular/efeitos da radiação , Proteínas de Ligação a DNA/deficiência , Detergentes , Humanos , Antígeno Nuclear de Célula em Proliferação/metabolismo , Solubilidade , Fatores de Tempo , Proteína de Xeroderma Pigmentoso Grupo ARESUMO
Recent mutagenesis studies of the ion channel proteins have allowed us to identify amino acid residues critical in determining ion selectivity. Ion selectivity of a channel can be altered even by single amino acid substitutions. Functional analyses of mutants largely support views in classical biophysics that the pore size and the fixed charges are major determinants of ion selectivity. For full understanding of the molecular mechanism of ion selectivity, elucidation of the tertiary structure of channel proteins remains essential.
Assuntos
Canais Iônicos/química , Canais Iônicos/fisiologia , Sequência de Aminoácidos , Ativação do Canal Iônico , Canais Iônicos/genética , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Receptores de Glutamato/química , Receptores de Glutamato/genética , Receptores de Glutamato/fisiologia , Receptores Nicotínicos/química , Receptores Nicotínicos/genética , Receptores Nicotínicos/fisiologia , Relação Estrutura-AtividadeRESUMO
The complete amino acid sequence of a novel ryanodine receptor/calcium release channel from rabbit brain has been deduced by cloning and sequence analysis of the cDNA. This protein is composed of 4872 amino acids and shares characteristic structural features with the skeletal muscle and cardiac ryanodine receptors. RNA blot hybridization analysis shows that the brain ryanodine receptor is abundantly expressed in corpus striatum, thalamus and hippocampus, whereas the cardiac ryanodine receptor is more uniformly expressed in the brain. The brain ryanodine receptor gene is transcribed also in smooth muscle.
Assuntos
Encéfalo/metabolismo , Canais de Cálcio/química , Cálcio/metabolismo , Receptores Colinérgicos/química , Sequência de Aminoácidos , Animais , Aorta , Sequência de Bases , Canais de Cálcio/genética , Canais de Cálcio/metabolismo , Clonagem Molecular , DNA , Dados de Sequência Molecular , Músculos/metabolismo , Miocárdio/metabolismo , Especificidade de Órgãos/genética , Coelhos , Receptores Colinérgicos/genética , Receptores Colinérgicos/metabolismo , Canal de Liberação de Cálcio do Receptor de Rianodina , Homologia de Sequência de AminoácidosRESUMO
The cardiac dihydropyridine-sensitive L-type calcium channel was stably expressed in Chinese hamster ovary cells by transfecting the rabbit cardiac calcium channel alpha 1 subunit cDNA with or without coexpression of the beta subunit of skeletal muscle calcium channel. Whereas coexpression of the beta subunit significantly increased DHP binding activity and calcium channel activity, it did not affect the amount of the alpha 1 subunit expressed, as judged by RNA blot hybridization analysis and immunoblotting analysis. The results suggest that association with the beta subunit is necessary for the alpha 1 subunit protein to take a proper conformation suitable for a functional calcium channel.
Assuntos
Canais de Cálcio/química , Animais , Células CHO , Canais de Cálcio/fisiologia , Cricetinae , Expressão Gênica , Técnicas In Vitro , Substâncias Macromoleculares , Miocárdio/química , Conformação Proteica , Coelhos , Proteínas Recombinantes , Relação Estrutura-AtividadeRESUMO
The channel pore of the nicotinic acetylcholine receptor (AChR) has been investigated by analysing single-channel conductances of systematically mutated Torpedo receptors expressed in Xenopus oocytes. The mutations mainly alter the size and polarity of uncharged polar amino acid residues of the acetylcholine receptor subunits positioned between the cytoplasmic ring and the extracellular ring. From the results obtained, we conclude that a ring of uncharged polar residues comprising threonine 244 of the alpha-subunit (alpha T244), beta S250, gamma T253 and delta S258 (referred to as the central ring) and the anionic intermediate ring, which are adjacent to each other in the assumed alpha-helical configuration of the M2-containing transmembrane segment, together form a narrow channel constriction of short length, located close to the cytoplasmic side of the membrane. Our results also suggest that individual subunits, particularly the gamma-subunit, are asymmetrically positioned at the channel constriction.
Assuntos
Canais Iônicos/genética , Oócitos/fisiologia , Receptores Nicotínicos/genética , Sequência de Aminoácidos , Animais , Feminino , Canais Iônicos/fisiologia , Substâncias Macromoleculares , Potenciais da Membrana , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Receptores Nicotínicos/fisiologia , Proteínas Recombinantes/metabolismo , Torpedo , Xenopus laevisRESUMO
The SS2 and adjacent regions of the 4 internal repeats of sodium channel II were subjected to single mutations involving, mainly, charged amino acid residues. These sodium channel mutants, expressed in Xenopus oocytes by microinjection of cDNA-derived mRNAs, were tested for sensitivity to tetrodotoxin and saxitoxin and for single-channel conductance. The results obtained show that mutations involving 2 clusters of predominantly negatively charged residues, located at equivalent positions in the SS2 segment of the 4 repeats, strongly reduce toxin sensitivity, whereas mutations of adjacent residues exert much smaller or no effects. This suggests that the 2 clusters of residues, probably forming ring structures, take part in the extracellular mouth and/or the pore wall of the sodium channel. This view is further supported by our finding that all mutations reducing net negative charge in these amino acid clusters cause a marked decrease in single-channel conductance.
Assuntos
Mapeamento de Peptídeos , Saxitoxina/genética , Canais de Sódio/química , Tetrodotoxina/genética , Potenciais de Ação/efeitos dos fármacos , Sequência de Aminoácidos , Animais , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Ratos , Saxitoxina/farmacologia , Canais de Sódio/efeitos dos fármacos , Tetrodotoxina/farmacologiaRESUMO
Glutamic acid residues in the SS2 segment of the internal repeats III and IV of the brain calcium channel BI were subjected to single point mutations. The mutant channels were tested for macroscopic current properties and sensitivities to inorganic blockers. The mutation that replaces glutamic acid 1,469 with glutamine altered ion-selection properties and strongly reduced the sensitivity to Cd2+, whereas the analogous mutation of glutamic acid 1,765 exerted smaller effects on ion-selection properties. Our results indicate that these glutamic acid residues, equivalently positioned in the aligned sequences, play different roles in the selective permeability of the calcium channel.
Assuntos
Encéfalo/fisiologia , Canais de Cálcio/fisiologia , Sequência de Aminoácidos , Animais , Técnicas In Vitro , Ativação do Canal Iônico , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , Coelhos , Sequências Repetitivas de Ácido Nucleico , Relação Estrutura-Atividade , Xenopus laevisRESUMO
Cloning and sequence analysis of cDNA showed that the brain type of ryanodine receptor (RYR) is expressed in human Jurkat T-lymphocyte cells. Fura-2 measurements revealed that the RYR in T-cells functions as a ryanodine-sensitive, caffeine-insensitive Ca2+ release channel. Furthermore, ryanodine stimulated proliferation and altered the growth pattern of cultured human T-cells when added together with FK506.
Assuntos
Canais de Cálcio/metabolismo , Proteínas Musculares/metabolismo , Linfócitos T/citologia , Sequência de Aminoácidos , Sequência de Bases , Química Encefálica , Cafeína/farmacologia , Cálcio/metabolismo , Canais de Cálcio/genética , Divisão Celular/efeitos dos fármacos , Clonagem Molecular , DNA Complementar/análise , Expressão Gênica , Humanos , Dados de Sequência Molecular , Proteínas Musculares/genética , RNA Mensageiro/análise , Rianodina/farmacologia , Canal de Liberação de Cálcio do Receptor de Rianodina , Alinhamento de Sequência , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos , Linfócitos T/efeitos dos fármacos , Tacrolimo/farmacologia , Células Tumorais CultivadasRESUMO
The complete amino acid sequences of two potassium channel proteins from NG108-15 neuroblastoma-glioma hybrid cells have been deduced by cloning and sequencing the cDNAs. One of these proteins (NGK2) is structurally more closely related to the Drosophila Shaw gene product than to the Shaker and Shab gene products, whereas the other (NGK1) is identical with a rat brain potassium channel protein (BK2) which is more closely related to the Drosophila Shaker gene product. mRNAs derived from both the cloned cDNAs, when injected into Xenopus oocytes, direct the formation of functional potassium channels with properties of delayed rectifiers.
Assuntos
Canais de Potássio/fisiologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Clonagem Molecular , DNA/genética , Camundongos , Dados de Sequência Molecular , Filogenia , Ratos , Xenopus laevisRESUMO
We have cloned and sequenced the cDNA of the human brain ryanodine receptor (RyR3), which is composed of 4866 amino acids and shares characteristic structural features with the rabbit RyR3. Northern blot analysis shows that the human RyR3 mRNA is abundantly expressed in hippocampus, caudate nucleus and amygdala as well as in skeletal muscle. The human RyR3 mRNA is also detected in several cell lines derived from human brain tumors. Functional expression of RyR3 and a chimeric RyR suggests that RyR3 forms a calcium-release channel with a very low Ca2+ sensitivity.
Assuntos
Encéfalo/metabolismo , Canal de Liberação de Cálcio do Receptor de Rianodina/metabolismo , Animais , Clonagem Molecular , DNA Complementar , Humanos , Coelhos , Canal de Liberação de Cálcio do Receptor de Rianodina/genética , Análise de Sequência , Análise de Sequência de DNA , Distribuição Tecidual , Células Tumorais CultivadasRESUMO
We studied a 59-year-old woman with dementia, retinitis pigmentosa, sensorimotor neuropathy, and attacks of vomiting associated with blood pressure lability and loss of consciousness. Abnormalities included CT evidence of cerebral atrophy, low IQ, slow central and peripheral nerve conduction velocities, axonal degeneration in sural nerve biopsy, and elevated levels of catecholamines and slow waves in EEG during attacks. Her sister, two brothers, and daughter also had progressive muscle weakness, visual disturbance, and similar vomiting attacks. The hereditary nervous system disorder does not fit any previously described condition.
Assuntos
Demência/complicações , Disautonomia Familiar/complicações , Retinose Pigmentar/complicações , Adulto , Biópsia , Eletromiografia , Potenciais Somatossensoriais Evocados , Feminino , Neuropatias Hereditárias Sensoriais e Autônomas , Humanos , Masculino , Pessoa de Meia-Idade , Condução Nervosa , Linhagem , Nervo Sural/patologia , Nervo Sural/ultraestrutura , Tomografia Computadorizada por Raios XRESUMO
To gain an insight into the molecular basis of ion permeation mechanism through the nicotinic acetylcholine receptor (AChR) channel, we have determined permeability ratios of organic cations relative to Na+ of specifically mutated Torpedo californica AChR channels expressed in Xenopus oocytes. The mutations involved mainly the side chains of the amino acid residues in the intermediate ring, where mutations have been found to exert strong effects on single-channel conductance and ion selectivity among alkali metal cations. The results obtained reveal that both the size and the net charge of the side chains of the intermediate ring are involved in determining the permeability, and provide experimental evidence that the pore size at the intermediate ring is a critical determinant of permeability. Our findings further suggest that changes in net charge exert effects on permeability by affecting the pore size of the channel.