Distribution of parvalbumin, calretinin, and calbindin-D28k immunoreactivity in the rat amygdaloid complex and colocalization with ?-aminobutyric acid

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Abstract
To understand the organization of inhibitory circuitries in the rat amygdala, the distribution of parvalbumin, calretinin, and calbindin immunoreactivity was investigated in the rat amygdaloid complex. Colocalization of various calcium-binding proteins with the inhibitory transmitter gamma-aminobutyric acid (GABA) was studied by using the mirror technique. Parvalbumin-immunoreactive (-ir) elements were located mostly in the deep amygdaloid nuclei, whereas the calretinin-ir and calbindin-ir staining were most intense in the cortical nuclei as well as in the central nucleus and the amygdalohippocampal area. Second, the distribution of immunopositive neurons largely parallelled the distribution of terminal and neuropil labeling. Third, immunostained neurons could be divided into four major morphologic types (types 1-4) based on the characteristics of the somata and the dendritic trees. The fourth lightly stained neuronal type that had a pyramidal GABA-negative soma was observed only in calretinin and calbindin preparations. Fourth, parvalbumin-ir terminals formed basket-like plexus and cartridges, which suggests that parvalbumin labels GABAergic inhibitory basket cells and axo-axonic chandelier cells, respectively. Colocalization studies indicated that 521 of 553 (94%) of parvalbumin-ir, 419 of 557 (75%) of calbindin-ir, and 158 of 657 (24%) of calretinin-ir neurons were GABA-positive in the deep amygdaloid nuclei. A high density of large GABA-negative calbindin-ir neurons was observed caudally in the medial division of the lateral nucleus and GABA-negative calretinin-ir neurons were observed in the magnocellular division of the accessory basal nucleus as well as in the intermediate and parvicellular divisions of the basal nucleus. These data suggest that in various amygdaloid areas, neuronal excitability is controlled by GABAergic neurons that contain different calcium-binding proteins. The appearance of basket-like plexus and cartridges in the parvalbumin preparations, but not in calretinin preparations, suggests that like in the hippocampus, the distribution of inhibitory terminals in the dendritic and perisomatic regions of postsynaptic neurons in the rat amygdala is organized in a topographic manner.