Sanders D, Hansen U P, Slayman C L
Proc Natl Acad Sci U S A. 1981 Sep;78(9):5903-7. doi: 10.1073/pnas.78.9.5903.
Possible methods by which eukaryotic cells can regulate intracellular pH (pHi) in response to experimental acid loading were investigated by using as a model cell the fungus Neurospora. Attention was focused on the role of membrane transport in such regulation, starting from the fact that this organism possesses a powerful electrogenic proton extrusion pump. Intracellular acidification was forced by introducing butyric acid into the recording medium, and subsequent changes in pHi and membrane potential were determined with intracellular microelectrodes. In separate experiments, membrane current-voltage curves were obtained and resolved--by an explicit kinetic model--into distinct pump and leak components. Decreased pHi causes increased outward pumping of H+ ions, in a manner quantitatively consistent with their role as a substrate for the proton pump. This increased pumping is often manifest as a transient hyperpolarization at the onset of cytoplasmic acidification. With a considerably slower time course, decreased pHi also produces a large increase in membrane leak conductance, which brings about net membrane depolarization and further stimulates the pump (by virtue of the reduced back electromotive force). Although the identity of the ion responsible for increased leak conductance is not yet known, the evident modulation of conductance seemingly plays an important role in stabilizing the intracellular pH: Stimulation of the pump alone would have little net effect on pHi because it would result simply in enhanced backflux of H+ (to which the membrane is most permeable in normal circumstances). An increased leak to nonprotons, however, would allow the pump to accomplish net H+ ejection.
以真菌粗糙脉孢菌作为模型细胞，研究了真核细胞响应实验性酸负荷调节细胞内pH（pHi）的可能机制。研究重点是膜转运在这种调节中的作用，研究起始于该生物体拥有强大的生电质子外排泵这一事实。通过将丁酸引入记录介质来强制细胞内酸化，并用细胞内微电极测定随后pHi和膜电位的变化。在单独的实验中，获得膜电流-电压曲线，并通过一个明确的动力学模型将其解析为不同的泵电流和漏电流成分。pHi降低会导致H⁺离子向外泵出增加，其方式在数量上与它们作为质子泵底物的作用一致。这种增加的泵出通常表现为细胞质酸化开始时的短暂超极化。在相当缓慢的时间进程中，pHi降低还会使膜漏导大幅增加，这会导致膜去极化，并进一步刺激泵（由于反向电动势降低）。尽管尚不清楚导致漏导增加的离子身份，但电导的明显调节似乎在稳定细胞内pH方面起着重要作用：仅刺激泵对pHi几乎没有净效应，因为这只会导致H⁺反向通量增加（在正常情况下膜对H⁺的通透性最高）。然而，对非质子的漏导增加将使泵能够实现净H⁺排出。
