Effect of guanine nucleotides on the dark voltage of single frog rods

Abstract

Single frog rods consisting of the outer segment and the ellipsoid were investigated by the whole-cell patch-clamp technique. When the recording pipette was filled with a simple intracellular medium containing potassium as the principal cation, a slow increase in dark voltage (hyperpolarization) associated with a decay of the photoresponses was observed. The hyperpolarization started at a dark voltage of -27 ± 8 mV, followed an exponential course, and leveled out at -52 ± 6 mV. The time constant was proportional to the access resistance of the preparations. With a pipette medium containing a 0.5 or 1.0 /iM cGMP, the initial dark voltage was shifted to more positive values and the tendency of hyperpolarization was clearly attenuated. Similar results were obtained with 1 mM GTP. The effects of GDP and of ATP were less significant. In experiments with 1 mM GTP plus 1 mM ATP, the dark voltage behaved as in experiments with only GTP. The stabilizing action of GTP was amplified by EGTA so that with 1 mM GTP plus 1 mM free EGTA the dark voltage was stable at a level of -15 mV. It is concluded that the preparations lose intracellular components such as cGMP and GTP by diffusion into the recording pipette and that the losses are prevented or reduced when the pipette medium contains these nucleotides in nearly physiological concentrations. For the internal transmitter cGMP, the results suggest that its free concentration does not exceed 1 ^M.

Sodium nitroprusside alters dark voltage and light responses in isolated retinal rods during whole-cell recording

“…When the recording pipette was filled with a medium devoid of nucleotides, the dark voltage of the cells hyperpolarized during the recording period from an initial value of about -25 mV to more than -5 0 mV (Schmidt et al, 1989a). This hyperpolarization can be attributed to a loss of nucleotides by diffusion from the cytoplasm into the recording pipette.…”

“…This loss is indicated by a slow hyperpolarization following an exponential course with time constant T (Schmidt et al, 1989a). This loss is indicated by a slow hyperpolarization following an exponential course with time constant T (Schmidt et al, 1989a).…”

Effect of intracellularly applied sodium ions on the dark voltage of isolated retinal rods

“…The data of this study reflect a diffusional exchange of compounds between the cytoplasm of the rod and the interior of the recording pipette (see also Schmidt et al, 1989). Under standard conditions, the cells behave as if they lose internal components of relevance for the normal dark current or dark voltage.…”

“…Frogs (Rana esculenta) were dark-adapted and the eyes were enucleated after decapitation. Rod outer segments with attached ellipsoids were isolated as described before (Schmidt et al, 1989). The incubating medium had the following composition: NaCl, 100 mM; KC1, 2.7 mM; MgCl 2 , 0.5 mM; CaCl 2) 1 mM; glucose, 5 mM; HEPES, 10 mM; pH 7.8.…”

“…Previously, we investigated this system in single isolated frog rods (Schmidt et al, 1989). The cells were patch-clamped and recorded in the whole-cell mode.…”