Use of confocal microscope for the cellular analysis of the glycine synaptic receptor

The presence of glycine receptors was examined with a monoclonal antibody and indirect immunofluorescence on reticular neurons of the goldfish (Carassius auratus) brainstem. Images of thin (0.6 microns) optical sections were recorded from 80 microns thick specimen with a confocal microscope thus obviating the need for mechanical slicing. Due to the reduced out-of-focus noise, high resolution was obtained. Lookthrough projections were computer generated. Compared with classical methods involving serial sectioning, this approach allowed the analysis of the subcellular distribution of this receptor with a considerable gain of time and increased resolution. On the Mauthner cell, an identified reticulo-spinal neuron, we found, that the size of glycine receptor microdomains varies depending on the cellular localization, i.e. somatic or dendritic. Furthermore the intensity of fluorescence was uneven within individual clusters, probably reflecting differences in receptor concentration. These heterogeneities may influence the variance of synaptic inhibitory noise in different regions of the Mauthner cell.