Syncytial isopotentiality, resulting from a strong electrical coupling, emerges as a physiological mechanism that coordinates individual astrocytes to function as a highly efficient system in brain homeostasis. However, whether syncytial isopotentiality occurs selectively to certain brain regions or is universal to astrocytic networks remains unknown. Here, we have explored the correlation of syncytial isopotentiality with different astrocyte subtypes in various brain regions. Using a nonphysiological K+‐free/Na+ electrode solution to depolarize a recorded astrocyte in situ, the existence of syncytial isopotentiality can be revealed: the recorded astrocyte's membrane potential remains at a quasi‐physiological level due to strong electrical coupling with neighboring astrocytes. Syncytial isopotentiality appears in Layer I of the motor, sensory, and visual cortical regions, where astrocytes are organized with comparable cell densities, interastrocytic distances, and the quantity of directly coupled neighbors. Second, though astrocytes vary in their cytoarchitecture in association with neuronal circuits from Layers I–VI, the established syncytial isopotentiality remains comparable among different layers in the visual cortex. Third, neurons and astrocytes are uniquely organized as barrels in Layer IV somatosensory cortex; interestingly, astrocytes both inside and outside of the barrels do electrically communicate with each other and also share syncytial isopotentiality. Fourth, syncytial isopotentiality appears in radial‐shaped Bergmann glia and velate astrocytes in the cerebellar cortex. Fifth, although fibrous astrocytes in white matter exhibit a distinct morphology, their network syncytial isopotentiality is comparable with protoplasmic astrocytes. Altogether, syncytial isopotentiality appears as a system‐wide electrical feature of astrocytic networks in the brain.

BackgroundNeonatal astrocytes are diverse in origin, and undergo dramatic change in gene expression, morphological differentiation and  syncytial networking throughout development. Neonatal astrocytes also play multifaceted roles in neuronal circuitry establishment. However, the extent to which neonatal astrocytes differ from their counterparts in the adult brain remains unknown.ResultsBased on ALDH1L1-eGFP expression or sulforhodamine 101 staining, neonatal astrocytes at postnatal day 1–3 can be reliably identified in hippocampal stratum radiatum. They exhibit a more negative resting membrane potential (VM), −85 mV, than mature astrocytes, −80 mV and a variably rectifying whole-cell current profile due to complex expression of voltage-gated outward transient K+ (IKa), delayed rectifying K+ (IKd) and inward K+ (IKin) conductances. Differing from NG2 glia, depolarization-induced inward Na+ currents (INa) could not be detected in neonatal astrocytes. A quasi-physiological VM of −69 mV was retained when inwardly rectifying Kir4.1 was inhibited by 100 μM Ba2+ in both wild type and TWIK-1/TREK-1 double gene knockout astrocytes, indicating expression of additional leak K+ channels yet unknown. In dual patch recording, electrical coupling was detected in 74 % (14/19 pairs) of neonatal astrocytes with largely variable coupling coefficients. The increasing gap junction coupling progressively masked the rectifying K+ conductances to account for an increasing number of linear voltage-to-current relationship passive astrocytes (PAs). Gap junction inhibition, by 100 μM meclofenamic acid, substantially reduced membrane conductance and converted all the neonatal PAs to variably rectifying astrocytes. The low density expression of leak K+ conductance in neonatal astrocytes corresponded  to a ~50 % less K+ uptake capacity compared to adult astrocytes.ConclusionsNeonatal astrocytes predominantly express a variety of rectifying K+ conductances, form discrete cell-to-cell gap junction coupling and are deficient in K+ homeostatic capacity.

We have recently shown that a linear current-to-voltage (I-V) relationship of membrane conductance (passive conductance) reflects the intrinsic property of K+ channels in mature astrocytes. While passive conductance is known to underpin a highly negative and stable membrane potential (VM) essential for the basic homeostatic function of astrocytes, a complete repertoire of the involved K+ channels remains elusive. TREK-1 two-pore domain K+ channel (K2P) is highly expressed in astrocytes, and covalent association of TREK-1 with TWIK-1, another highly expressed astrocytic K2P, has been reported as a mechanism underlying the trafficking of heterodimer TWIK-1/TREK-1 channel to the membrane and contributing to astrocyte passive conductance. To decipher the individual contribution of TREK-1 and address whether the appearance of passive conductance is conditional to the co-expression of TWIK-1/TREK-1 in astrocytes, TREK-1 single and TWIK-1/TREK-1 double gene knockout mice were used in the present study. The relative quantity of mRNA encoding other astrocyte K+ channels, such as Kir4.1, Kir5.1, and TREK-2, was not altered in these gene knockout mice. Whole-cell recording from hippocampal astrocytes in situ revealed no detectable changes in astrocyte passive conductance, VM, or membrane input resistance (Rin) in either kind of gene knockout mouse. Additionally, TREK-1 proteins were mainly located in the intracellular compartments of the hippocampus. Altogether, genetic deletion of TREK-1 alone or together with TWIK-1 produced no obvious alteration in the basic electrophysiological properties of hippocampal astrocytes. Thus, future research focusing on other K+ channels may shed light on this long-standing and important question in astrocyte physiology.