Objective To observe the electrophysiological and morphological features of retinal ganglion cells (RGCs) in rats, and investigate its effect on the visual signal conduction. Methods Whole cell recordings were obtained from 112 RGCs of 30 rats at the age of 7-30 days. Resting membrane potential (RMP) was recorded, and input impedance was noted after given 2 mV hyperpolarizing current by voltage clamp. The action potential (AP) was induced by deplorizing current at different densities. The histological staining was actualized by injecting with biotin into the RGCs, and the diameter of the cells was measured. Results Three different discharge patterns of RGCs in response to maintained depolarizing currents were recorded: single spike (25 RGCs), transient firing (40 RGCs), and sustained firing (47 RGCs).The dia meter was 14-16μm in 57.14% transient firing RGCs, and 10-12 μm in 62.50% sustained firing RGCs. The maximum frequency of AP of sustained firing RGCs was significantly higher than that of transient firing RGCs (P＜0.05). Conclusion The single firing of RGCs was an immature electrophysiological feature. The electrophysiological features of transient firing and sustained firing RGCs may be important to make the visual information code in spatial and temporal pathway. The electrophysiological and morphological features of RGCs in rats may be correlated with each other. (Chin J Ocul Fundus Dis，2004，20：160-164)
Objective To investigate the relationship between electrophysiological and morphological properties of neurons in visual cortex of developing rat, speculate the coincided degree between electrophysiological and morphological change and realize the mechanism of normal visual development. Methods Whole cell patch-clamp recording and intracellular staining were used to acquire cellular microelectrode recording in visual cortex from Sprague-Dawley rats (4~28 days old). The histological process was made. Results The differences of electrical feature between pyramidal cells and non-pyramidal cells were significant. The morphological maturity degree is different in developing visual cortex. Conclusion The different function of pyramidal and non-pyramidal cells in local integrition is reflected by their electrical feature in the process of visual development. In critical period of visual development, the coincision degree of the electrophysiological and morphological change in visual cortex is larger than that in the subcortex constructure. (Chin J Ocul Fundus Dis, 2001,17:289-292)
【摘要】 目的 探讨辣椒素对不同年龄SD大鼠内脏感觉神经元上辣椒素受体（TRPV1）介导的离子通道的影响。 方法 急性分离7~9 d和21~23 d大鼠迷走神经结状神经节神经元，利用全细胞膜片钳技术在分离的神经元上记录辣椒素激活TRPV1受体后通道电流的变化。 结果 ①7~9 d和21~23 d大鼠内脏感觉神经元的膜电容分别为（18.57±8.60）和（19.85±9.47） pF，（Pgt;0.05）；②辣椒素能够激活7~9 d和21~23 d大鼠内脏感觉神经元上TRPV1并产生相似的内向电流，两组产生的峰电流密度分别为（48.59±18.87）、（55.91±20.52） pA/pF（Pgt;0.05）；③反复应用辣椒素使TRPV1受体发生失敏现象。 结论 大鼠内脏感觉神经元的TRPV1受体通道在出生后已经发育成熟，且对辣椒素激活的通道电流有相似的变化。【Abstract】 Objective To investigate the effects of capsaicin on transient receptor potential vanilloid 1 (TRPV1) receptor-mediated ion channel currents of visceral sensory neurons in different-aged Sprague-Dawley rats. Methods We isolated the vagal nodose ganglion neurons of rats at an age of 7-9 days or 21-23 days acutely. With the whole cell patch clamp technique, we recorded the current changes of TRPV1 channels activated by capsaicin. Results ① Membrane capacitances of the visceral sensory neurons were (18.57±8.60) and (19.85±9.47) pF in rats of 7-9 and 21-23 days, respectively (Pgt;0.05). ② Capsaicin activated the TRPV1 channels and generated inward currents in all the rats; and the peak current densities of the rats of 7-9 days and 21-23 days were respectively (48.59±18.87) and (55.91±20.52) pA/pF (Pgt;0.05). ③ Repeated applications of capsaicin produced a phenomenon of desensitization in TRPV1 channels. Conclusion TRPV1 receptor channels of visceral sensory neurons in rats have matured after birth, and the current changes of TRPV1 channels activated by capsaicin are similar.
ObjectiveTo observe the changes in open probability and protein expression of large conductance Ca2+-activated K+ (BK) channel in retinal vascular smooth muscle cells (RVSMCs) of diabetic rats. MethodsStreptozotocin (STZ)-induced rat diabetic animal model was established by STZ injection intraperitoneally.RVSMCs were isolated by enzyme digestion. The BK currents in control and diabetic groups were recorded by patch clamp technique in single channel configuration. BK channel protein expression in control and diabetic group were measured by Western blot. ResultsCompared with control group, the NP0 of BK channels in diabetic group were significantly increased (t=4.260, P < 0.05). Compared with control group, there was no significant difference inα-subunit protein expression in diabetic group in RVSMCs (t=10.126, P > 0.05); however, β1-subunit protein expression was remarkably increased in diabetic group (t=5.146, P < 0.05). ConclusionThe NP0 of BK channels andβ1-subunit protein expression are increased in RVSMCs of diabetic rats.
Objective To investigate the effects of docosahexenoic acid (DHA) on large conductance Ca2+-activated K+ (BK) channels in normal retinal artery smooth muscle cells (RASMCs). Methods Cultured human RASMCs (6 th-8 th generations) were used to patch clamp experiment. The open probabihties (NP0) in BK channels with different concentrations (0.0, 1.0, 3.0, 5.0, 7.5, 10.0 μmol/L) of DHA were recorded by patch clamp technique in single channel configuration. RASMCs were intervened by different concentrations (0.0, 1.0, 5.0 μmol/L) of DHA as control group, low and high doses of DHA groups, respectively. The protein expressions of β subunit of BK channels in RASMCs from three groups were measured by Western blot. Results The NP0 of BK channels were 0.044 4±0.001 2, 0.081 2±0.004 2, 0.209 0±0.006 1, 0.310 5±0.005 3, 0.465 0±0.007 8 and 0.497 7±0.014 5 with perfusate of 0.0, 1.0, 3.0, 5.0, 7.5, 10.0 μmol/L DHA. DHA activated BK channels in a dose-dependent manner (F=2.621,P＜0.05). There was no significant difference in the protein expression of control group, low and high doses of DHA groups (F=11.657,P＞0.05). Conclusion DHA can directly activate BK channels, no increasing in subunit expression of BK channels.
Patch clamp is a technique that can measure weak current in the level of picoampere (pA). It has been widely used for cellular electrophysiological recording in fundamental medical researches, such as membrane potential and ion channel currents recording, etc. In order to obtain accurate measurement results, both the resistance and capacitance of the pipette are required to be compensated. Capacitance compensations are composed of slow and fast capacitance compensation. The slow compensation is determined by the lipid bilayer of cell membrane, and its magnitude usually ranges from a few picofarads (pF) to a few microfarads (μF), depending on the cell size. The fast capacitance is formed by the distributed capacitance of the glass pipette, wires and solution, mostly ranging in a few picofarads. After the pipette sucks the cells in the solution, the positions of the glass pipette and wire have been determined, and only taking once compensation for slow and fast capacitance will meet the recording requirements. However, when the study needs to deal with the temperature characteristics, it is still necessary to make a recognition on the temperature characteristic of the capacitance. We found that the time constant of fast capacitance discharge changed with increasing temperature of bath solution when we studied the photothermal effect on cell membrane by patch clamp. Based on this phenomenon, we proposed an equivalent circuit to calculate the temperature-dependent parameters. Experimental results showed that the fast capacitance increased in a positive rate of 0.04 pF/℃, while the pipette resistance decreased. The fine data analysis demonstrated that the temperature rises of bath solution determined the kinetics of the fast capacitance mainly by changing the inner solution resistance of the glass pipette. This result will provide a good reference for the fine temperature characteristic study related to cellular electrophysiology based on patch clamp technique.