Alkondon M, Albuquerque E X
Department of Pharmacology and Experimental Therapeutics, University of Maryland School of Medicine, Baltimore 21201.
J Recept Res. 1991;11(6):1001-21. doi: 10.3109/10799899109064693.
The properties of the neuronal nicotinic acetylcholine receptor in primary cultures of hippocampal cells from fetal rats (17-18 days gestation) were studied using the whole-cell patch-clamp technique in Na(+)-external, Cs(+)-internal and nominally Mg(2+)-free solutions. The nicotinic agonists acetylcholine, (+)anatoxin-a, and (-) and (+)nicotine all evoked inward whole-cell currents in hippocampal neurons that were voltage clamped near their resting potentials. Sensitivity to (+)anatoxin-a was first detected at around day 6, and thereafter the magnitude of the response increased as a function of number of days in culture up to about 40 days. The whole-cell current waveforms consisted of more than one peak whose relative amplitude depended on the agonist concentration. These currents were reversibly blocked by micromolar concentrations of d-tubocurarine, mecamylamine, and dihydro-beta-erythroidine. At nanomolar concentrations, neuronal bungarotoxin, alpha-bungarotoxin and alpha-cobratoxin caused an irreversible blockade of the currents but they were unaffected by tetrodotoxin, atropine, DL-2-amino-5-phosphonovaleric acid, Mg2+, and 6,7-dinitroquinoxaline-2,3-dione. In addition, the currents were also blocked in a reversible manner by methyllycaconitine at picomolar concentration. The current-voltage plots elicited by both (+)anatoxin-a and acetylcholine revealed larger inward currents and smaller or no outward currents. The present results demonstrate the existence of an inwardly rectifying, snake neurotoxin-sensitive functional nicotinic acetylcholine receptor ion channel in rat hippocampal neurons.
采用全细胞膜片钳技术，在细胞外为Na⁺、细胞内为Cs⁺且无Mg²⁺的溶液中，研究了胎鼠（妊娠17 - 18天）海马细胞原代培养物中神经元烟碱型乙酰胆碱受体的特性。烟碱型激动剂乙酰胆碱、（+）anatoxin - a、（ - ）和（+）尼古丁均可在电压钳制于静息电位附近的海马神经元中诱发内向全细胞电流。对（+）anatoxin - a的敏感性最早在第6天左右被检测到，此后，响应幅度随培养天数增加而增大，直至约40天。全细胞电流波形由多个峰组成，其相对幅度取决于激动剂浓度。这些电流可被微摩尔浓度的d - 筒箭毒碱、美加明和二氢β - 刺桐啶可逆性阻断。在纳摩尔浓度下，神经元型银环蛇毒素、α - 银环蛇毒素和α - 眼镜蛇毒素可导致电流的不可逆阻断，但它们不受河豚毒素、阿托品、DL - 2 - 氨基 - 5 - 膦酸戊酸、Mg²⁺和6,7 - 二硝基喹喔啉 - 2,3 - 二酮的影响。此外，皮摩尔浓度的甲基lycaconitine也能以可逆方式阻断电流。由（+）anatoxin - a和乙酰胆碱引发的电流 - 电压曲线显示，内向电流较大，外向电流较小或无外向电流。目前的结果表明，大鼠海马神经元中存在一种内向整流、对蛇神经毒素敏感的功能性烟碱型乙酰胆碱受体离子通道。
