Figure 3.

Electrophysiological and molecular analyses of pyramids and perivascular astrocytes. A, Infrared Dodt gradient contrast (IR-DGC) image of a layer III pyramidal cell that showed the typical firing of a regular spiking neuron with frequency adaptation and amplitude accommodation of action potentials in current clamp recordings (B). C, This pyramidal cell also expressed VGluT1 and COX-2; the high molecular weight band in the COX-2 lane corresponds to genomic amplification. RT-mPCR protocol performed on 500 pg of total cortical RNAs showing PCR products corresponding to the expect sizes for VGluT1, COX-1, and COX-2 amplicons. D, SR101 vital staining of perivascular cortical astrocytes. Wide-field fluorescence imaging of a SR101-labeled cortical slice (left panel) showing an intensely labeled astrocyte (arrow) and its process (arrowhead) in the vicinity of a diving arteriole (*). The pial surface is on the right side. Corresponding field under IR-DGC (middle panel); superimposition of the two images is shown in the right panel. The arrow indicates the SR101-stained astrocyte that was recorded in whole-cell configuration depicted in E. Note the linear I/V curve and the hyperpolarized resting membrane potential characteristic of passive astrocytes. The inset illustrates representative current responses evoked by voltage steps (from −180 mV to +40 mV, 20 mV increments) used to determine the I/V curve at steady state. F, RT-mPCR analysis of the SR101-positive astrocyte (shown in A and B) revealed expression of GFAP and S100β, but not COX-1 or COX-2, whereas total cortical RNAs showed PCR products of the expect size for all amplicons. Lanes Φ correspond to the molecular weight marker Phi X 174/HaeIII.