The Effect On Ion Channel Of Neuron By Low-Level Laser Irradiation And The Method To Current Detection

The research for bio-stimulation effect of low-level laser irradiation is in a relatively fundamental stage. And its mechanism is not still clarified. In this dissertation, a series of researches were taken from the viewpoint of excitability of cell membrane using the patch clamp technique, which focused on the bio-stimulation effect of low-level laser irradiation. The theory and experiments included the time dependency, frequency dependency, voltage dependency, reversibility, does-effect correlation, steady-state activation and inactivation kinetics of ion channel currents, action potential of neuron and its firings induced by low-level laser as well as the method to detection of ionic single-channel currents.The properties of voltage-gated Na+ and K+ channel of neuron induced by low-level laser irradiation were studied using the whole-cell patch clamp technique. The experimental results showed that low-level laser irradiation significantly affected the activation potentials and current amplitudes of ion channel. The amplitudes of INa, IA and IK reversibly changed in a time-dependent and voltage-dependent, but not frequency-dependent manner. In addition, the action of low-level laser showed itself irradiating-does dependency and correlation to energy density of laser. The kinetic characteristics of steady-state activation and inactivation altered and the curves of activation and inactivation were shifted by low-level laser irradiation. Therefore, low-level laser irradiation induced the changes of voltage-gated ionic channel property of neurons. It is suggested that the bio-stimulation effect of low-level laser was potentially correlative to the configuration and function of ion channel.The characteristics of action potential and its firings of neuron induced by low-level laser irradiation were investigated using the whole-cell current clamp technique. The experiments revealed that the repetitive firings of the normal neuron were elicited by continuous ramp currents and pulse currents respectively more than threshold intensity and increased with input excitatory current intensity. Under low-level laser irradiation, the resting potential and threshold potential of neurons lowered, action potential amplitude increased, action potential duration of 90% repolarization (APD₉₀) widened, firing frequency of action potential slowed. And the effects were mostly recovered after the end of laser irradiating. The results suggest that low-level laser irradiation could change the properties of action potential, which facilitated excitability of neurons and improved its conductive function.Because of the weakness of the single ionic channel current of cell membrane, the background noise always dominates in the patch-clamp recordings. The restoration of ionic single-channel signal based on HMM was an effective means of idealizing patch-clamp recordings. For the defects of weak classifying ability and the parameters falling into local maximum in HMM training, in this dissertation, a hybrid model of HMM and RBF network was applied to restore ionic single-channel currents under the white background noise. To ensure parameters'convergence to the maximal likelihood value, we used a global optimization algorithm of HMM parameters based on the stochastic relaxation. The experimental results of simulating and practical sampling data from patch-clamp system have shown the effectiveness and feasibility of the methods.The patch-clamp technique provided a new method to exploring bio-stimulation effect of low-level laser in cell and molecule level. Theory and experimental results disclosed by this dissertation establish a solid basis of the researches for the bio-stimulation mechanism of low-level laser and the microinformation detection to ionic single-channel currents.