Immunohistochemical Study on the Nitric Oxide Synthase Neurons in the Brain Stem of Rats

Na, Jun Hee; Huh, Young Buhm; Park, Chan; Lee, Won Kyu; Kim, Jung Hye; Yoo, Jin Hwa; Ahn, Hee Kyung

Abstract

Nitric oxide (NO), a free radical that has been postulated to act as a neurotransmitter, neuromodulator, or second messenger molecule in nervous system, is synthesized from L-arginine by nitric oxide synthase (NOS). The NADPH-diaphorase (NADPH-d) histochemical techenique provides a simple and robust method to stain the selected populations of NOS neurons in the brain. This study was aimed to clarify the distribution of NOS neurons in the brain stem of rats. To verify the distribution of NOS neurons in the brain stem, the neurons were stained by the NOS immunohis-tochemistry and NADPH-d histochemistry. Image analyzer-assisted densitometry and cell counting method have been used to quantitatively characterize groups of neuronal cells. Double labeling of NOS immunohistochemistry and NADPH-d histochemistry showed the coexistence of NOS and NADPH-d in same neurons. Neuronal cell bodies exhibiting NOS/NADPH-d staining were found in particular nuclei throughout the brain stem. The number of NOS/NADPH-d neurons were variable in brain stem nuclei. The NADPH-d neurons exhibited different intensities of reaction product. Some groups, including paradorsal raphe nucleus, laterodorsal tegmental nucleus and pedunculopontine tegmental nucleus were extremely heavily stained. Other neurons such as those in the interpeduncular nucleus, central gray, substantia nigra lateralis, nucleus solitarius and raphe obscurus nucleus were moderately stained, while other neurons such supragenual nucleus, lateral paragigantocellular nucleus and prepositus hypoglossal nucleus were weakly stained. The present study describes the many locations within the brain stem in which NADPH-d occurs. Since NADPH-d activity colocalizes with NOS, the results indicate the likely involvement of nitric oxide in the neuronal functions of many brain stem nuclei of rats.