ABSTRACT
In order to identify whether functional L-type calcium channels are expressed in neural stem cells(NSCs) from rat embryonic hippocampus, and whether L-type calcium channels participate in the modulation of proliferation and differentiation of NSCs, the rat embryonic hippocampal tissue was dispersed into a single cell suspension, and the dissociated cells were cultured in serum-free DMEM/F12 medium containing epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), N-2 and B27 supplement. Immunofluorescent labeling showed an expression of nestin-positive cells. Following five-days culture in differentiation medium, neuron-like and astrocyte-like cells were observed, which expressed ?-tubulin Ⅲ(Tuj1) and Glial fibrillary acidic protein (GFAP), respectively. Western blot analysis showed an expression of Cav1.2?1C subunits in NSCs, but no Cav1.3?1D subunits. Moreover, L-type calcium channel currents were recorded in those cells by using whole-cell patch clamp techniques. It was found that activation of L-type calcium channels promotes proliferation and differentiation to neuronal type of NSCs. The results indicated that rat embryonic hippocampal NSCs express functional L-type calcium channels, L-type calcium channels modulate proliferation and differentiation of NSCs.
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Objective To study the effect of S2-protein from SARS coronavirus on the chloride channel currents in A549 cells and its possible cellular mechanisms. Methods The chloride channel currents were recorded in cultured A549 cells by using the whole-cell mode of patch clamp techniques. The experiments were divided into four groups: Control group: chloride channel currents were recorded in untreated A549 cells; S2 protein group: currents were recorded in A549 cells treated with S2 protein (final concentration 50?g/ml); calphostin C + S2 protein group: the effect of S2 protein on the currents in A549 cells pretreated with calphostin C (0.1mmol/L) for 10 minutes; SB203580+S2 protein group: the effect of S2 protein on the currents was examined with the solution containing SB203580 (20?mol/L). Results The currents of chloride channel in normal A549 cells showed outwardly rectifying properties and were insensitive to both TEA and amiloride, but were significantly inhibited both by SITS and DIDS (P
ABSTRACT
Objective Effects of aqueous extract from Lishi No.5 formula on concentration of intracellular free-calcium fluorescent intensity in neurons. Methods Hippocampus neurons of 1-day newborn SD rats were cultured with conventional culture technique. The cells cultured for 9-12 days were used for experiment. Intracellular free calcium fluorescent intensity of neurons cultured under different conditions was assayed with confocal microscopic calcium image technique after loading of Fluo-3/AM. Results Free-calcium concentration was enhanced by aqueous extract from Lishi No.5 formula, this concentration was up to 126.35?9.35nmol/L, but the concentration is at normal scope; L-type calcium channel blocker Nifedipine may block the effect of aqueous extract from Lishi No.5 formula enhancing intracellular free-calcium concentration in part, it make intracellular free-calcium concentration down to 90.75?10.15nmol/L, but Nifedipine itself also decreased markedly free-calcium concentration to 40.65?5.65nmol/L. NMDA increase Markedly calcium fluorescent intensity, the effects of NMDA was decreased notably after pre-treating with aqueous extract from Lishi No.5 formula. The effect of MK-801, an antagonist of NMDA receptor, on inhibition of NMDA increasing free calcium fluorescent intensity was significantly reduced after pretreatment of aqueous extract from Lishi No.5 formula. Conclusion Aqueous extract from Lishi No.5 formula maybe bidirectional adjust intracellular free calcium concentration via enhancing L-type calcium channel activity and blocking NMDA receptor in part.
ABSTRACT
Objective:To explore the role of reactive oxygen species(ROS,i.e,H_2O_2 and O_2-) in regulation of respiratory rhythm in the medial area of nucleus retrofacialis(mNRF).Methods: Medullary slices of neonatal SD rats,including hypoglossal nerve(Ⅻn) and mNRF,were made according to Suzue's method.Simultaneous recording of the Ⅻn respiratory rhythmic activity(RRA) with suction electrode and the respiratory neuronal discharge were performed with whole cell patch in the mNRF on the brainstem slice in vitro.The effect of t-butyl hydroperoxide(tBHP) and ?-lipoic acid(?-LA) on the respiratory pacemaker neurons and respiratory rhythm in the mNRF were observed.Results: tBHP significantly decreased respiratory cycle(RC) and increased respiratory amplitude;?-LA significantly increased RC and decreased its amplitude.Meanwhile,?-LA significantly prolonged the action potential of the respiratory cadmium-insensitive pacemaker neurons and reduced its amplitude,but it had no significant effect on the cadmium-sensitive respiratory pacemaker neurons.Voltage steps and ramps showed that ?-LA inhibited both the transient and persistent sodium current of cadmium-insensitive pacemaker neurons.Conclusion: ROS has an excitatory effect on the respiratory rhythm and the cadmium-insensitive respiratory pacemaker neurons through modulating sodium current.