Spinal cord astrocytes in vitro: Phenotypic diversity and sodium channel immunoreactivity

Abstract

The expression of sodium channels in morphologically and antigenically distinct astrocytes derived from neonatal rat spinal cords was examined at various times in culture. During the course of this study [2-40 days in vitro (DIV)], nine morphologies of glial fibrillary acidic protein (GFAP)+ cells were distinguished: 1a) flat, fibroblast-like; 1b) elongated, with generally few, short processes; 1c) triangular soma with three short, stubby processes; 1d) bipolar with long, slender processes; 1e) bipolar with broad, flared processes; 1f) stellate with radially oriented slender processes extending from a small to moderate-sized soma; 1g) multiple short, stubby processes extending from a moderate-sized soma; 1h) flat, roundish shape with either a smooth edge ("pancake"-like) or numerous very short processes; and 1i) broad, elongated cell body with orthogonally oriented short, spike-like processes. Not all cell types were present at all times in culture. Each type of astrocyte displayed sodium channel immunoreactivity at some time in culture; however, different types of astrocytes exhibited different patterns, over time, of sodium channel staining. Sodium channel immunoreactivity in all astrocyte types was reduced to low levels by 14 DIV, and was not detectable at 40 DIV. Except for types 1b and 1e, A2B5 staining was present on all astrocyte morphologies at some time in culture, and was generally attenuated with longer times in vitro; in contrast to cultures derived from neonatal rat optic nerve, A2B5 staining does not distinguish unequivocally between the various classes of morphologically different astrocytes derived from spinal cord. O4 immunoreactivity was consistently observed only on bipolar, elongated, and process-bearing astrocytes, though not all process-bearing astrocytes were O4+. These results demonstrate that astrocytes derived from neonatal spinal cord are morphologically and antigenically heterogeneous. Moreover, while spinal cord astrocytes express sodium channels, these astrocytes exhibit a time-course of channel expression that is different from astrocytes derived from several other CNS regions where sodium channel staining is maintained even for extended times in culture, suggesting a regional modulation of astrocyte function.

Differential modulation of TTX-sensitive and TTX-resistant Na+ channels in spinal cord astrocytes following activation of protein kinase C

“…1) (Miller and Szigeti, 1991;Black et al, 1993). Na+ currents in stellate cells express TTX-S Na+ currents (& = 5.7 nM) with kinetic features similar to most cultured neurons.…”

Region-specific astrogliosis in brains of mice heterozygous for mutations in the neurofibromatosis type 1 (Nf1) tumor suppressor

“…Astrocytes in specific brain regions differ (e.g., Ref. [3]); astrocyte differences could account for the sensitivity of these regions to effects of loss of NF1. In contrast to our results in Nf1/nf1 mice, human brains analyzed showed widespread astrogliosis [39].…”

Sodium Channel Expression: A Dynamic Process in Neurons and Non-Neuronal Cells

“…Sodium channel III mRNA and protein is also expressed in both stellate and fiat astrocytes at low levels ( fig. 3e, 0-The expression of sodium currents and sodium channels II and III in cultured spinal cord astro cytes is not invariant; rather, the level of expression in both fiat and stellate astrocytes changes with time in cul ture [8,85]. The expression of sodium channel protein isoforms in fiat and stellate astrocytes with increasing time in culture is shown in table 1.…”

“…Astrocytes cultured from P-0 rat spinal cords have previously been shown to be phenotypically diverse [8,63]; however, at 5 -7 DIV, two astrocyte morphologies are predominant: flat, fibroblast-like and stellate, pro cess-bearing, together comprising 80-90% of the GFAP-positive cells present in these cultures. In spinal cord cultures, stellate, process-bearing astrocytes express a 'neuronal-like' type of sodium current, while flat, 'pancake'-like astrocytes express 'glial-like' sodium currents in which voltage dependence is shifted in hyperpolarizing direction [85,87], Astrocytes derived from P-0 rat spinal cords and examined for rat brain type sodium channels exhibit con sistent patterns of expression when probed with isotypespecific riboprobes in in situ hybridization studies ( fig.…”