Regulation of intracellular pH in salamander retinal rods.

Abstract

1. We measured intracellular pH (pHi) in rods isolated from the retina of the axolotl salamander, Ambystoma mexicanum, using the fluorescent indicator 2',7'-bis(carboxyethyl)-5(and -6)-carboxyfluorescein (BCECF).2. The light exposures associated with data acquisition had no marked effect on pH,. There was no sharp change between the value obtained from the first exposure of dark-adapted rods and subsequent readings. Increasing the acquisition frequency from 1 to 10 min-' either had no effect, or brought about a slow acidification, which was stopped or reversed when the low frequency was restored.3. In nominally HCO3--free solution at pH 7 5, the rods had a steady-state pHi of 7'09 + 0-02 (n = 46) and a buffering power (,Bi) of 24 + 1 mm (pH unit)f' (n = 48). The buffering power was virtually constant in the pH range 6-6-8-0. In the same range, pHi depended linearly on perfusion pH (pHO) with regression coefficients of 0 4-0 5. 7. In the presence of CO2-HC03-, 100 ,m 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS) decreased the acid extrusion rate by 20 % on average. Lowering the extracellular Clconcentration to 7 mm raised pH,, but did not significantly affect the acid extrusion rate.8. We conclude that retinal rods regulate pHi by both Na+-H+ exchange and mechanism(s)involving HC03--Cl-exchange. In the present conditions, the Na+-H+ exchanger appears as the dominant mechanism for acid extrusion.Phototransduction in vertebrate photoreceptor cells is sensitive to pH. Early work showed that the amplitude of mass rod responses from the frog retina dropped by 70% when perfusion pH (pH.) was decreased from 8 to 6 (Sillman,

Intracellular pH modulates inner segment calcium homeostasis in vertebrate photoreceptors

“…We conclude that an increase in pH i modulates Ca 2ϩ release from internal stores in rod perikarya, possibly by regulating the open probability of ryanodine channels (2,18,20,79). In contrast to the well-understood pH sensitivity of voltage-operated channels (3,52,67,59,70), PMCAs (61,72), and Ca 2ϩ stores (2,79), little is known about pH i sensitivity of neuronal SOCE. Our results regarding photoreceptor SOCE are inconclusive.…”

“…This calculation is similar to the 50 mM/(pH unit) Ϫ1 obtained by Liebman et al (45) for toad rod outer segments and higher than ϳ24 mM/(pH unit) Ϫ1 calculated previously for salamander rod outer segments (59). This estimate is also higher than values reported for rat Purkinje cells [10.7 mM/(pH unit) Ϫ1 ; 25] and cortical neurons [27 mM/(pH unit) Ϫ1 ; 35] but roughly equivalent to 60 mM/(pH unit) Ϫ1 measured in cardiomyocytes (74 -75).…”

“…Hence, HEPES has typically been the buffer of choice in studies that characterized pH, chloride, and calcium signaling mechanisms in isolated photoreceptors (3-4, 17, 34, 37, 39 -41, 50, 55, 60, 63-66, 68 -69) and horizontal cells (67) and has also been used in investigations of synaptic transmission at photoreceptor synapses (28,37). In the present study, the main mechanism regulating cytosolic acid loads under nominally bicarbonatefree conditions was the ubiquitous electroneutral Na ϩ /H ϩ antiporter, which couples proton transport to the sodium gradient (34,59). Vertebrate photoreceptors may also express electroneutral Cl Ϫ /HCO 3 Ϫ exchange (28,34,38,59) and electrogenic Na ϩ -HCO 3 Ϫ cotransport and Na ϩ -driven C Ϫ /HCO 3 Ϫ exchange mechanisms (10,83), which could be functionally coupled to carbonic anhydrases within the bicarbonate transport metabolon (62).…”

“…Intracellular acidity is often measured in terms of pH i [the negative logarithm, base 10, of the molar concentration of dissolved protons or hydronium ions (H 3 O ϩ )]; a decrease in pH i thus indicates an increase in intracellular H ϩ concentration ([H ϩ ] i ). Intracellular pH (pH i ) and intracellular Ca 2ϩ concentration ([Ca 2ϩ ] i ) shifts are not independent of each other: while pH i modulates the affinity of intracellular Ca 2ϩ -binding sensor proteins and activity of most known Ca 2ϩ channels and transporters (5, 19 -20, 30, 32-33, 36, 61, 70 -72), changes in [Ca 2ϩ ] i and calmodulin (CaM) activity cause pH i shifts through modulation of Ca 2ϩ /H ϩ and/or Na ϩ /H ϩ exchange mechanisms (44,51,61,67,72).Activity-dependent pH i and [Ca 2ϩ ] i changes are especially pronounced in metabolically active cells, such as vertebrate photoreceptors (26,29,38,59,80). Light and darkness are associated with intracellular production and export of protons (11,21,26,(37)(38).…”

“…Activity-dependent pH i and [Ca 2ϩ ] i changes are especially pronounced in metabolically active cells, such as vertebrate photoreceptors (26,29,38,59,80). Light and darkness are associated with intracellular production and export of protons (11,21,26,(37)(38).…”