Colocalization of calcium-binding proteins and GABA in neurons of the rat basolateral amygdala

Abstract

The basolateral amygdala contains subpopulations of non-pyramidal neurons that express the calcium-binding proteins parvalbumin, calbindin-D28k (calbindin) or calretinin. Although little is known about the exact functions of these proteins, they have provided useful markers of specific neuronal subpopulations in studies of the neuronal circuitry of the cerebral cortex and other brain regions. The purpose of the present study was to investigate whether basolateral amygdalar non-pyramidal neurons containing parvalbumin, calbindin, or calretinin exhibit immunoreactivity for GABA, and to determine if calretinin is colocalized with parvalbumin or calbindin in the rat basolateral amygdala. Pyramidal neurons were distinguished from non-pyramidal neurons on the basis of staining intensity. Using immunofluorescence confocal laser scanning microscopy, as well as the ‘mirror technique’ on immunoperoxidase-stained sections, it was found that there was virtually no colocalization of calretinin with parvalbumin or calbindin, but that the great majority of basolateral amygdalar non-pyramidal neurons containing parvalbumin, calbindin, or calretinin exhibited GABA immunoreactivity. Calbindin-positive neurons constituted almost 60% of the GABA-containing population in both subdivisions of the basolateral nucleus and more than 40% of the GABA-containing population in the lateral nucleus. Parvalbumin-positive neurons constituted 19–43% of GABA-immunoreactive neurons in the basolateral amygdala, depending on the nucleus. Calretinin-positive non-pyramidal neurons constituted about 20% of the GABA-positive neuronal population in each nucleus of the basolateral amygdala. These findings indicate that non-pyramidal neurons containing parvalbumin, calbindin, or calretinin comprise the majority of GABA-containing neurons in the basolateral amygdala, and that the calretinin subpopulation is distinct from non-pyramidal subpopulations containing parvalbumin and calbindin. These separate neuronal populations may play unique roles in the inhibitory circuitry of the amygdala.