Distribution of NADPH-diaphorase and expression of nNOS, N-methyl-D-aspartate receptor (NMDAR1) and non-NMDA glutamate receptor (GlutR2) genes in the neurons of the hippocampus after domoic acid-induced lesions in adult rats.
Neuronal degeneration followed by detection of nitric oxide (NO)-producing neurons of the hippocampus was investigated at 4 h, 16 h, 24 h, 2 days, 5 days, and 14 days after administration of domoic acid (DA), in the present study. Histopathological analysis (Nissl staining) displayed dark-stained degenerating neurons in the hippocampus at 24 h to 14 days after DA administration, with degeneration most severe at 5-14 days. NADPH-d-positive neurons were observed in different subfields of the hippocampus in control rats and DA treated rats at 4-24 h. Complete loss of NADPH-d-positive neurons in the CA1 and CA3 subfields and also in the hilus of dentate gyrus (DG) was observed at 5 days and 14 days after the administration of DA. In contrast, at 4-24 h, neuronal nitric oxide synthase (nNOS)-immunoreactive cells were absent from the hippocampal subfields in control and DA-treated animals but were observed at 5 days and 14 days after DA administration. N-methyl-D-aspartate receptor (NMDAR1) immunoreactivity was increased in the hippocampal neurons at 5 days after DA administration and double immunofluorescence demonstrated its coexpression with induced nNOS expression. No significant change could be observed in the immunoreactivity of non-NMDA receptor (GlutR2) as compared with the controls, while occasional immunoreactive neurons were colocalized with induced nNOS expression. Reverse transcription-polymerase chain reaction analysis showed the upregulated expression of nNOS and downregulated expression of NMDAR1 at 5 days after the administration of DA. Although nNOS mRNA expression was rapidly induced at 5 days after DA administration, in situ hybridization analysis revealed complete loss of nNOS mRNA expression in the region of neuronal degeneration in the hippocampus at 24 h and 5 days after DA administration. The present study has shown that NADPH-d and nNOS express differentially in the neurons of the hippocampus in DA-induced neurotoxicity. It is speculated that induction of nNOS and glutamate receptor genes in the neurons of the hippocampus in response to DA-induced neurotoxicity could have contributed to the neuronal degeneration.
Ananth
C
C
Department of Anatomy, Faculty of Medicine, National University of Singapore, Singapore.
Dheen
S Thameem
ST
Gopalakrishnakone
P
P
Kaur
C
C
D000818:Animals; D004396:Coloring Agents; D019084:Fluorescent Antibody Technique, Indirect; D006624:Hippocampus / Q000166:cytology / Q000201:enzymology / Q000378:metabolism*; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D007608:Kainic Acid / Q000031:analogs & derivatives* / Q000633:toxicity*; D008297:Male; D018613:Microscopy, Confocal; D009252:NADPH Dehydrogenase / Q000378:metabolism*; D009467:Neuromuscular Depolarizing Agents / Q000633:toxicity*; D009474:Neurons / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000096:biosynthesis*; D052248:Nitric Oxide Synthase Type I; D051381:Rats; D017208:Rats, Wistar; D018094:Receptors, Metabotropic Glutamate / Q000096:biosynthesis / Q000235:genetics; D016194:Receptors, N-Methyl-D-Aspartate / Q000096:biosynthesis* / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D004396:Coloring Agents; D007608:Kainic Acid; D009252:NADPH Dehydrogenase; D009467:Neuromuscular Depolarizing Agents; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I; D018094:Receptors, Metabotropic Glutamate; D016194:Receptors, N-Methyl-D-Aspartate; C108822:metabotropic glutamate receptor 2; C496322:Nos1 protein, rat; C012301:domoic acid