Heterogeneity of Astrocyte Resting Membrane Potentials and Intercellular Coupling Revealed by Whole-Cell and Gramicidin-Perforated Patch Recordings from Cultured Neocortical and Hippocampal Slice Astrocytes

Abstract

Astrocytes are thought to regulate the extracellular potassium concentration by mechanisms involving both voltage-dependent and transport-mediated ion fluxes combined with intercellular communication via gap junctions. Mechanisms regulating resting membrane potential (RMP) play a fundamental role in determining glial contribution to buffering of extracellular potassium and uptake of potentially toxic neurotransmitters. We have investigated the passive electrophysiological properties of cultured neocortical astrocytes and astrocytes recorded in hippocampal slices from 18-25 d postnatal rats. These experiments revealed a wide range of astrocyte RMPs that were independent of developmental factors, length of culturing, cellular morphology, the electrophysiological techniques used (whole-cell vs perforated recording), cell-specific expression of Na ϩ /2HCO 3 Ϫ co-transporters, or voltage-dependent Na ϩ channels. Exposure of cultured astrocytes to differentiation-inducing factors (such as cAMP) or inhibition of proliferation (by serum deprivation) did not significantly influence RMP. Expression of ATP-sensitive potassium channels was absent in these glia; thus, K (ATP) -related mechanisms did not contribute to cell resting potential. In both cultured and slice astrocytes, spontaneous electrophysiological changes were commonly observed. These reversible events, which resulted in differential sensitivity to potassium channel blockers (cesium and barium) and sudden current-voltage profile changes, were attributable to dynamic changes in cell-to-cell coupling, as confirmed by recordings from isolated pairs of cells. We conclude that the heterogeneity of astrocytic RMP and intercellular coupling both in culture and in situ are intrinsic properties of glia that may contribute to transcellular transport of potassium. We propose a model in which spatial buffering may be facilitated by heterogeneous mechanisms controlling glial RMP in combination with dynamic changes in intercellular coupling.

Do Glia Have Heart? Expression and Functional Role forEther-A-Go-GoCurrents in Hippocampal Astrocytes

“…For example, although most glial recordings reveal expression of inward rectifier currents (K IR ), it is not clear how spatial buffering consisting of accumulation and release of potassium may be mediated by exclusively inward potassium fluxes. Furthermore, the spatial buffering theory is based on the assumption that glial resting membrane potential (RMP) is close to E K , an assumption frequently challenged by direct experimental evidence (McKhann et al, 1997b). Recordings from astrocytes have demonstrated that although many have an RMP consistent with K IR expression, a significant proportion of neocortical and hippocampal astrocytes have RMPs more positive than E K , and hence, this is inconsistent with an exclusive role of K IR in determining RMP.…”

“…For example (1) while cloned K IR conductance is rather insensitive to [Na ϩ ] out , currents underlying inward rectification in glia are dramatically reduced in low sodium (Ransom and Sontheimer, 1995;Ransom et al, 1996); (2) in "complex" glia (D'Ambrosio et al, 1998), currents evoked by hyperpolarization are characterized by time-dependent activation/inactivation, a behavior that has not been described for K IR expressed in oocytes or in cardiac myocytes; and (3) RMP in glia does not always follow E K , suggesting that currents other than K IR are involved in RMP determination. The latter issue is discussed in detail elsewhere (McKhann et al, 1997b). Briefly, resting membrane potentials of Ϫ80 mV for in situ or cultured astrocytes have been commonly reported, but these measurements were biased by an exclusion criterion.…”

“…Briefly, resting membrane potentials of Ϫ80 mV for in situ or cultured astrocytes have been commonly reported, but these measurements were biased by an exclusion criterion. When exclusion criteria were not used, both hyperpolarized and relatively depolarized RMPs were measured (McKhann et al, 1997b). In a previous work, we demonstrated that both depolarized and hyperpolarized recordings were obtained from cells with astrocytic properties, as determined by electron microscopy of cells injected with biocytin during whole-cell recordings (D'Ambrosio et al, 1998).…”

“…Voltage-dependent currents were evoked from cultured or in situ astrocytes by using the whole-cell variation of the patchclamp technique McKhann et al, 1997b;D'Ambrosio et al, 1998). ERG currents are enhanced in solutions containing elevated extracellular potassium (Sanguinetti et al, 1995), hence membrane currents were studied at nonphysiological potassium concentrations in cultured astrocytes (40 mM, with NaCl decreased proportionally to 106 mM).…”

Carbenoxolone modifies spontaneous inhibitory and excitatory synaptic transmission in rat somatosensory cortex

“…GJs have been shown to exist between neocortical glial cells and have been implicated in spatial buffering of extracellular K + [22]. Uncoupling of glial cell GJs could lead to a rise in extracellular K + and, in turn, depolarization of neurons and heightened pyramidal cell excitability.…”