Analysis of K⁺ accumulation reveals privileged extracellular region in the vicinity of glial cells in situ

Abstract

Astrocytes and oligodendrocytes in rat and mouse spinal cord slices, characterized by passive membrane currents during de- and hyperpolarizing stimulation pulses, express a high resting K+ conductance. In contrast to the case for astrocytes, a depolarizing prepulse in oligodendrocytes produces a significant shift of reversal potential (Vrev) to positive values, arising from the larger accumulation of K+ in the vicinity of the oligodendrocyte membrane. As a result, oligodendrocytes express large tail currents (Itail) after a depolarizing prepulse due to the shift of K+ into the cell. In the present study, we used a mathematical model to calculate the volume of the extracellular space (ECS) in the vicinity of astrocytes and oligodendrocytes (ESVv), defined as the volume available for K+ accumulation during membrane depolarization. A mathematical analysis of membrane currents revealed no differences between glial cells from mouse (n = 59) or rat (n = 60) spinal cord slices. We found that the Vrev of a cell after a depolarizing pulse increases with increasing Itail, expressed as the ratio of the integral inward current (Qin) after the depolarizing pulse to the total integral outward current (Qout) during the pulse. In astrocytes with small Itail and Vrev ranging from -50 to -70 mV, the Qin was only 3-19% of Qout, whereas, in oligodendrocytes with large Itail and Vrev between -20 and 0 mV, Qin/Qout was 30-75%. On the other hand, ESVv decreased with increasing values of Vrev. In astrocytes, ESVv ranged from 2 to 50 microm3, and, in oligodendrocytes, it ranged from 0.1 to 2.0 microm3. Cell swelling evoked by the application of hypotonic solution shifted Vrev to more positive values by 17.2 +/- 1.8 mV and was accompanied by a decrease in ESVv of 3.6 +/- 1.3 microm3. Our mathematical analysis reveals a 10-100 times smaller region of the extracellular space available for K+ accumulation during cell depolarization in the vicinity of oligodendrocytes than in the vicinity of astrocytes. The presence of such privileged regions around cells in the CNS may affect the accumulation and diffusion of other neuroactive substances and alter communication between cells in the CNS.

Glia Contribute to the Purinergic Modulation of Inspiratory Rhythm-Generating Networks

“…Lack of a detectable ATP response in half of the nonexcitable cells supports heterogeneity of glia (Grass et al, 2004), but does not necessarily indicate that cells are insensitive to ATP, only that P2R stimulation did not evoke a change in membrane current. It is also possible that the ATP-insensitive cells are oligodendrocytes, as these also have high input resistance values (Chvátal et al, 2004;Káradó ttir et al, 2008). However, our data do not allow unequivocal discrimination because at the ages studied here (P0 -P4) astrocyte precursor cells, astrocytes, oligodendrocyte precursor cells, and oligodendrocytes all fit within the size range of recorded cells.…”

Diffusion in Brain Extracellular Space

“…The above-mentioned results prompted a quantitative analysis to determine the actual volumes of local ECS around these respective cell types (65). Based on a model (92, 308) (see Section II.H) incorporating K + fluxes, Chvátal and colleagues estimated that the width of the ECS around the cell bodies of oligodendrocytes was only 0.3 – 10 nm whereas the width around the cell bodies of astrocytes was 4 – 154 nm.…”

High extracellular K+ evokes changes in voltage-dependent K+ and Na+ currents and volume regulation in astrocytes

“…The reversal potential of the tail current (V rev tail ) was determined from the I/V relationship measured 5 ms after the offset of a 20 ms depolarizing (+20 mV) prepulse followed by a series of 20-ms de-and hyperpolarizing pulses ranging from −130 to +20 mV. The K + accumulation in the vicinity of the cell membrane after a depolarizing prepulse (ΔK + ) was determined by subtracting the values of the extracellular K + concentration before ( K þ m  à e ) and after the depolarizing prepulse ( K þ rev  à e ) [7,8]. Because the astrocytic membrane is almost exclusively permeable to K + , K þ m  à e and K þ rev  à e can be calculated using the Nernst equation from the corresponding values of V m and V rev tail , respectively, i.e.,…”

“…Complex astrocytes (b) are characterized by the presence of delayed outwardly rectifying K + (K DR ), A-type K + (K A ), inwardly rectifying K + (K IR ), and Na + currents (I Na ); passive astrocytes (d) show mainly passive conductance and additional voltage-dependent K + currents, while astrocyte precursors (f) are characterized by the presence of K DR , K A , and I Na , but no K IR and negligible passive conductance effect of high K + on astrocyte volume changes evoked by hypotonic stress. Previously, we have shown that spinal cord astrocytes in rats and mice display no significant interspecies differences in cell swelling or membrane properties when exposed to hypotonic stress [2,7,44]. Therefore, we have taken advantage of the direct visualization of astrocytes in spinal cord slices from EGFP/GFAP mice to quantify the differences in volume changes between astrocytes in control tissue and those from spinal cords incubated in ACF50.…”

“…The exposure of astrocytes to elevated [K + ] e results in reversible membrane depolarization, the accumulation of intracellular K + , and rapid cell swelling [30,47]. Astrocyte swelling, which can be induced in vitro by hypotonic solution or by an isotonic solution with an increased [K + ] [2,8,17], evokes a large accumulation of K + in the vicinity of the cell membrane after a depolarizing prepulse, resulting from an extracellular space (ECS) volume decrease around swollen astrocytes [7,8,44]. Sykova et al [43] showed that incubating rat spinal cords in 50 mM K + evokes cell swelling, resulting in a decrease in the ECS volume fraction and astrocyte activation [manifested as an increase in glial fibrillary acidic protein (GFAP) immunoreactivity].…”