ATP-mediated increase in H⁺ flux from retinal Müller cells: a role for Na⁺/H⁺ exchange

Abstract

Small alterations in extracellular H+ can profoundly alter neurotransmitter release by neurons. We examined mechanisms by which extracellular ATP induces an extracellular H+ flux from Müller glial cells, which surround synaptic connections throughout the vertebrate retina. Müller glia were isolated from tiger salamander retinae and H+ fluxes examined using self-referencing H+-selective microelectrodes. Experiments were performed in 1 mM HEPES with no bicarbonate present. Replacement of extracellular sodium by choline decreased H+ efflux induced by 10 µM ATP by 75%. ATP-induced H+ efflux was also reduced by Na+/H+ exchange inhibitors. Amiloride reduced H+ efflux initiated by 10 µM ATP by 60%, while 10 µM cariporide decreased by 37%, and 25 µM zoniporide reduced H+ flux by 32%. ATP-induced H+ fluxes were not significantly altered by the K+/H+ pump blockers SCH28080 or TAK438, and replacement of all extracellular chloride with gluconate was without effect on H+ fluxes. Recordings of ATP-induced H+ efflux from cells simultaneously whole-cell voltage-clamped revealed no effect of membrane potential from -70 mV to 0 mV. Restoration of extracellular potassium after cells were bathed in 0 mM potassium produced a transient alteration in ATP-dependent H+ efflux. The transient response to extracellular potassium occurred only when extracellular sodium was present and was abolished by 1 mM ouabain, suggesting alterations in sodium gradients mediated by Na+/K ATPase activity. Our data indicate that the majority of H+ efflux elicited by extracellular ATP from isolated Müller cells is mediated by Na+/H+ exchange.

Extracellular ATP-Induced Alterations in Extracellular H+ Fluxes From Cultured Cortical and Hippocampal Astrocytes

“…Recent work examining the molecular mechanisms underlying ATP-induced H + extrusion from retinal Müller glia suggests that the majority of acid is provided by Na + /H + exchange activity (Tchernookova et al, 2021). Consequently, we sought to determine if this was also the case for the ATP-induced extrusion of H + from brain astrocytes.…”

“…The lack of dependence on extracellular sodium and the inability of Na + /H + inhibitors to reduce the flux differ from the ATP-induced alteration in H + flux previously detected in the radial glial cells (Müller cells) isolated from the retina of the tiger salamander. Upon removal of extracellular sodium, ATP-initiated H + fluxes from Müller cells were reduced by about 70%, and ATP-initiated H + fluxes were also inhibited by several pharmacological agents known to interfere with Na + /H + exchange (Tchernookova et al, 2021). These differences suggest that the molecular mechanism mediating the majority of H + flux in astrocytes cells is likely to be different from that in the retinal radial glial cells.…”

Review and Hypothesis: A Potential Common Link Between Glial Cells, Calcium Changes, Modulation of Synaptic Transmission, Spreading Depression, Migraine, and Epilepsy—H+

“…Additional experiments demonstrated that about 75% of the ATP-elicited increase in H + flux likely resulted from the extrusion of protons via Na + /H + exchange (Tchernookova et al, 2021). The complete removal of extracellular sodium and then with chelerythrine removed but ATP still present.…”

“…The experiments described above were carried out in solutions in which the primary extracellular H + buffer was 1 mM HEPES, with no bicarbonate added. ATP-elicited increases in H + flux from Müller cells also persisted unchanged when solutions were bubbled for 15 min before use with 100% oxygen, to remove trace amounts of CO 2 from the atmosphere that could potentially be used by very high affinity bicarbonate transporters (Theparambil et al, 2014;Tchernookova et al, 2021).…”

“…A potent but commonly overlooked regulator of synaptic transmission is simple H + -that is, small changes in levels of extracellular acidity around sites of neurotransmitter release. Recent studies have shown that activation of glial cells inducing increased intracellular calcium also promotes the release of H + from glia, and it has been proposed that this may play a key role in regulating synaptic transmission (Tchernookova et al, 2018(Tchernookova et al, , 2021Choi et al, 2021). In this review and hypothesis article, we first review studies demonstrating the potency of extracellular H + as a modulator of synaptic transmission and then describe techniques used and studies conducted to show calcium-dependent extrusion of H + from glial cells activated by extracellular ATP.…”