Distribution of calbindin-D28k, neuronal nitric oxide synthase, and nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d) in the lateral nucleus of the sheep amygdaloid complex

Abstract

This study describes calbindin-D28k (CB), neuronal nitric oxide synthase (nNOS), and nicotinamide adenine dinucleotide phosphate diaphorase (NADPH-d) expression in the lateral nucleus of the sheep amygdaloid complex. Double immunofluorescence protocol was used in order to determine whether there is colocalization of CB and nNOS. The CB-immunoreactive (IR) neuronal population was composed especially of non-pyramidal neurons, but a few pyramidal cells were also present. The non-pyramidal neurons showed a multipolar and, occasionally, a fusiform morphology. The comparison between single-labeled CB-IR non-pyramidal neurons and cells belonging to CB-IR neuronal population showed they were identical for morphology, mean size, and distribution. The single-labeled CB-IR non-pyramidal neurons were only the 17.8% of the total non-pyramidal neurons counted. The nNOS-IR neuronal population was represented by non-pyramidal multipolar and fusiform neurons. Single-labeled nNOS-IR non-pyramidal neurons had the same morphology, mean area, and distribution as cells belonging to nNOS-IR neuronal population. Single-labeled nNOS-IR non-pyramidal neurons were more numerous than single-labeled CB-IR, and represented the 73.7% of total non-pyramidal neurons counted. NADPH-d-positive cells had the same morphology and distribution as the nNOS-IR neurons. Double immunolabeling (CB/nNOS) was found mostly in non-pyramidal multipolar neurons and only in a few non-pyramidal fusiform cells. These neurons had a mean perikaryal area significantly higher and significantly smaller than that of single-labeled nNOS and single-labeled CB-IR non-pyramidal neurons, respectively. CB and nNOS coexist only in a minority of non-pyramidal neurons (8.5%). The 32.4% of all CB-IR non-pyramidal neurons were nNOS-positive; only 10.4% of nNOS-IR non-pyramidal neurons were CB-positive. These results indicate that CB and nNOS are expressed by selective neurons and that the majority of nNOS-IR non-pyramidal neurons are lacking in CB.