Relationships Between Calcium and pH in the Regulation of the Slow Afterhyperpolarization in Cultured Rat Hippocampal Neurons

Abstract

The Ca2+-dependent slow afterhyperpolarization (AHP) is an important determinant of neuronal excitability. Although it is established that modest changes in extracellular pH (pHo) modulate the slow AHP, the relative contributions of changes in the priming Ca2+ signal and intracellular pH (pHi) to this effect remain poorly defined. To gain a better understanding of the modulation of the slow AHP by changes in pHo, we performed simultaneous recordings of intracellular free calcium concentration ([Ca2+]i), pHi, and the slow AHP in cultured rat hippocampal neurons coloaded with the Ca2+- and pH-sensitive fluorophores fura-2 and SNARF-5F, respectively, and whole cell patch-clamped using the perforated patch technique. Decreasing pHo from 7.2 to 6.5 lowered pHi, reduced the magnitude of depolarization-evoked [Ca2+]i transients, and inhibited the subsequent slow AHP; opposite effects were observed when pHo was increased from 7.2 to 7.5. Although decreases and increases in pHi (at a constant pHo) reduced and augmented, respectively, the slow AHP in the absence of marked changes in preceding [Ca2+]i transients, the inhibition of the slow AHP by decreases in pHo was correlated with low pHo-dependent reductions in [Ca2+]i transients rather than the decreases in pHi that accompanied the decreases in pHo. In contrast, high pHo-induced increases in the slow AHP were correlated with the accompanying increases in pHi rather than high pHo-dependent increases in [Ca2+]i transients. The results indicate that changes in pHo modulate the slow AHP in a manner that depends on the direction of the pHo change and substantiate a role for changes in pHi in modulating the slow AHP during changes in pHo.

The slow afterhyperpolarization modulates high pH‐induced changes in the excitability of rat CA1 pyramidal neurons

“…The larger relative inhibition of the test response compared with the conditioning response seen at pH o 7.7 compared with pH o 7.4 could reflect the effect of high pH o to increase the neuronal response to the conditioning stimulus, thereby increasing the subsequent sAHP. However, we have recently reported (Kelly & Church, 2004, 2006) that high pH o ‐induced increases in s I ahp and the sAHP in hippocampal CA1 neurons are largely independent of increases in Ca 2+ entry through the HVA Ca 2+ channels that are the primary source of Ca 2+ for the activation of the sAHP in this cell type. These findings raise the possibility that the high pH o ‐induced increase in the sAHP evoked by an antidromic conditioning stimulus might reflect a more direct effect of high pH o to augment the sAHP independent of an increase in excitability to the conditioning stimulus.…”

“…Thus, the amplitude of the sAHP increased with the number of action potentials in the stimulus train (Fig. 2A and B) and decreased with membrane hyperpolarization (in four cells, the sAHP evoked by a train of three action potentials declined from 3.7 ± 0.3 mV at −60 mV to 1.9 ± 0.5 mV at −70 mV; P < 0.05; see Gustafsson & Wigström, 1981; Lancaster & Batchelor, 2000; Shah & Haylett, 2000; Wu et al ., 2004; Kelly & Church, 2006). In addition, the sAHP evoked by a single antidromic stimulus or by a train of three antidromic stimuli was markedly inhibited by the β‐adrenergic receptor agonist isoproterenol (8 µ m ) and the muscarinic receptor agonist carbachol (5 µ m ; Fig.…”

“…We have shown previously that the augmented s I ahp and sAHP observed in hippocampal neurons at high pH o remain sensitive to manoeuvres that modulate the current/potential at pH o 7.4 (Kelly & Church, 2004, 2006). Therefore, to examine whether the augmented sAHP observed after a single conditioning stimulus contributes to the marked inhibition of the test response compared with the conditioning response at pH o 7.7, we applied isoproterenol and Ba 2+ to modulate the sAHP.…”

“…These activity‐induced pH o shifts can in turn modulate the events that initially caused them, in part by affecting the activities of ligand‐ and voltage‐gated ion channels. Increases in pH o , for example, inhibit and augment, respectively, the activities of γ‐aminobutyric acid (GABA A ) and N ‐methyl‐ d ‐aspartate (NMDA) receptor‐operated channels, and increase Ca 2+ influx via high voltage‐activated (HVA) Ca 2+ channels (reviewed by Tombaugh & Somjen, 1998; Traynelis, 1998; see also Church et al ., 1998; Kelly & Church, 2004, 2006; Makani & Chesler, 2007). In turn, these effects contribute to the development of neuronal burst‐firing and epileptiform activity observed during extracellular alkalosis (Aram & Lodge, 1987; Balestrino & Somjen, 1988; Church & McLennan, 1989; see also Schuchmann et al ., 2006).…”

“…Suppression of the sAHP and its underlying current, s I AHP , transforms short bursts of action potentials into more prolonged discharges and increases the frequency of epileptiform events (Chamberlin & Dingledine, 1989; Jurgens et al ., 2005; Fernández de Sevilla et al ., 2006). Interestingly, however, increases in s I ahp and the sAHP accompany the increases in neuronal excitability and epileptiform activity induced by increases in pH o (Church & McLennan, 1989; see also Kelly & Church, 2004, 2006). This raises the question as to whether the augmented sAHP observed during increases in pH o may function to limit high pH o ‐induced increases in neuronal excitability, in a manner analogous to the action of the sAHP under basal conditions.…”