Electrical coupling of astrocytes in rat hippocampal slices under physiological and simulated ischemic conditions

Abstract

Mammalian protoplasmic astrocytes are extensively coupled through gap junction channels but the biophysical properties of these channels under physiological and ischemic conditions in situ are not well defined. Using confocal morphometric analysis of biocytin-filled astrocytic syncytia in rat hippocampal CA1 stratum radiatum we found that each astrocyte directly couples, on average, to 11 other astrocytes with a mean interastrocytic distance of 45 microm. Voltage-independent and bidirectional transjunctional currents were always measured between directly coupled astrocyte pairs in dual voltage-clamp recordings, but never from astrocyte-NG2 glia or astrocyte-interneuron pairs. The electrical coupling ratio varied considerably among astrocytes in developing postnatal day 14 rats (P14, 0.5-12.4%, mean = 3.6%), but became more constant in young adult P21 rats (0.18-3.9%, mean = 1.6%), and the coupling ratio declined exponentially with increasing pair distance. Electrical coupling was not affected by short-term oxygen-glucose deprivation (OGD) treatment, but showed delayed inhibition in an acidic extracellular pH of 6.4. Combination of acidic pH (6.4) and OGD, a condition that better represents cerebral ischemia in vivo, accelerated the inhibition of electrical coupling. Our results show that, under physiological conditions, 20.7-24.2% of K(+) induced currents can travel from any astrocytic soma in CA1 stratum radiatum to the gap junctions of the nearest neighbor astrocytes, but this should be severely inhibited as a consequence of the OGD and acidosis seen in the ischemic brain.

In Vivo Labeling of Cortical Astrocytes with Sulforhodamine 101 (SR101)

“…Additionally, SR101 staining appears to depend on animal age: Although the vast majority of astrocytes in slices prepared from animals 15 d or older are SR101 positive, only a fraction of cells with glial morphology are stained in younger animals . This developmental dependence of SR101 staining appears plausible given the pronounced structural and functional changes astrocytes undergo during postnatal development (Bushong et al 2004;Xu et al 2009). Further, electrophysiological and optical studies in acute brain slices indicate that SR101 staining (1 µM; 20 min incubation) does not interfere with astrocyte physiology ) and that astrocytes stained by SR101 in juvenile and adult animals represent classical astrocytes (i.e., cells with passive membrane properties, glutamate transporter activity, and a high rate of gap junction coupling) ).…”

“…What is the precise mechanism responsible for SR101 uptake? Are reactive astrocytes that typically show reduced gap junction coupling (Xu et al 2009) also labeled by SR101? How can the differences in SR101 staining between rodent cortex and other central nervous system regions or species be reconciled (Keifer et al 1992;Ehinger et al 1994;Lavallee and Pflieger 2009)?…”

Gap junctions mediate intercellular spread of sodium between hippocampal astrocytes in situ

“…Our results now demonstrate that, in addition and primarily independent of a concomitant calcium wave, sodium signals evoked in a single cell in situ rapidly propagate to neighboring cells through gap junctions. Astrocytes in the stratum radiatum of rat hippocampus are directly coupled to on average 11 other astrocytes with a mean interastrocytic distance of 45 lm (Xu et al, 2010). Speed of sodium propagation was > 60 lm/s to the nearest neighbors, indicating that somatic sodium signals generated in a single cell will reach about 10 directly coupled astrocytes appreciably faster than classical calcium waves (Scemes and Giaume, 2006).…”

Effects of meclofenamic acid on limbic epileptogenesis in mice kindling models

“…This discrepancy might be due to the nonselective or indirect effects of MFA in interfering amgydaloid kindling acquisition. MFA's action on blocking both astrocytic and neuronal gap junction is unequivocal [1,3,6,21,30]. Accumulating evidence indicates that gap junctions may be involved in epileptogenesis.…”

“…Meclofenamic acid (MFA) is a fenamate that has also been used to block gap junction channels through both astrocytic Cx43 [6,21,30] and neuronal Cx36 [3,25]. Although MFA can inhibit seizures in tetanus toxin-induced refractory focal cortical epilepsy in rats [14], there are no further studies to test whether or not MFA has antiepileptogenic or disease-modifying effects in epileptogenic models.…”