Imaging of L-type Ca 2+ channels in olfactory bulb neurones using fluorescent dihydropyridine and a styryl dye

Abstract

We have imaged the fluorescence of dihydropyridine-Bodipy (fDHP) in cultured olfactory bulb neurones in order to investigate the subcellular distribution of L-type calcium channels in these neurones. The neurones were stained with both fDHP and the voltage-sensitive styryl dye RH414. The fluorescence emission maxima of these dyes were in the green and red ranges of the spectrum, respectively, and were recorded by the 2 photomultiplier channels of a laser scanning microscope. The fDHP images were ratioed with the RH414 images taken simultaneously. The resulting ratio images revealed the spatial distribution of the surface density of L-type calcium channels. This density was highest on somata, in particular at the base of dendrites, and decreased with the distance from this maximum. Two classes of dendritic L-type channel distributions were observed: one with a homogeneously low density and another one with a characteristic gradient of the L-type channel density along proximal dendrites. This subcellular localization of L-type calcium channels is discussed in the light of specific functional roles.