Kelly Tony, Church John
Department of Cellular and Physiological Sciences, University of British Columbia, 2177 Wesbrook Mall, Vancouver, BC V6T 1Z3, Canada.
Pflugers Arch. 2005 Dec;451(3):418-27. doi: 10.1007/s00424-005-1483-6. Epub 2005 Jul 27.
The weak bases NH(3) and trimethylamine (TMeA), applied externally, are widely used to investigate the effects of increasing intracellular pH (pH(i)) on neuronal function. However, potential effects of the compounds independent from increases in pH(i) are not usually considered. In whole-cell patch-clamp recordings from rat CA1 pyramidal neurons, bath application of 1-40 mM NH(4)Cl or TMeA HCl reduced resting membrane potential and input resistance, inhibited the medium and slow afterhyperpolarizations (AHPs) and their respective underlying currents, mI(ahp) and sI(ahp), and led to the development of depolarizing current-evoked burst firing. Examined in the presence of 1 microM TTX and 5 mM TEA with 10 mM Hepes in the recording pipette, NH(3) and TMeA increased pH(i) and the magnitudes of depolarization-evoked intracellular [Ca(2+)] transients, Ca(2+)-dependent depolarizing potentials, and inward Ca(2+) currents but reduced the slow AHP and sI(ahp). When internal H(+) buffering power was raised by including 100 mM tricine in the patch pipette, the effects of NH(3) and TMeA to increase pH(i) and augment Ca(2+) influx were attenuated whereas the reductions in the slow AHP and sI(ahp) (as well as membrane potential and input resistance) were maintained. The findings indicate that increases in pH(i) contribute to the increases in Ca(2+) influx observed in the presence of NH(3) and TMeA but not to the reductions in membrane potential, input resistance or the magnitudes of AHPs. The results have implications for the interpretation of data from experiments in which pH(i) is manipulated by the external application of NH(3) or TMeA.
外部施加的弱碱氨（NH₃）和三甲胺（TMeA）被广泛用于研究细胞内pH值（pH(i)）升高对神经元功能的影响。然而，通常未考虑这些化合物独立于pH(i)升高的潜在影响。在大鼠CA1锥体神经元的全细胞膜片钳记录中，浴槽中施加1 - 40 mM氯化铵（NH₄Cl）或三甲胺盐酸盐（TMeA HCl）会降低静息膜电位和输入电阻，抑制中等和慢速超极化后电位（AHPs）及其各自的基础电流，即mI(ahp)和sI(ahp)，并导致去极化电流诱发的爆发式放电。在记录电极中存在1 μM河豚毒素（TTX）、5 mM四乙铵（TEA）和10 mM Hepes的情况下进行检测，NH₃和TMeA会升高pH(i)以及去极化诱发的细胞内[Ca²⁺]瞬变、Ca²⁺依赖性去极化电位和内向Ca²⁺电流的幅度，但会降低慢速AHP和sI(ahp)。当通过在膜片钳电极中加入100 mM三羟甲基氨基甲烷（tricine）来提高内部H⁺缓冲能力时，NH₃和TMeA升高pH(i)和增加Ca²⁺内流的作用减弱，而慢速AHP和sI(ahp)（以及膜电位和输入电阻）的降低则得以维持。这些发现表明，pH(i)的升高促成了在NH₃和TMeA存在时观察到的Ca²⁺内流增加，但对膜电位、输入电阻或AHPs幅度的降低没有作用。这些结果对于解释通过外部施加NH₃或TMeA来操纵pH(i)的实验数据具有启示意义。
