In vivo inhibition of hippocampal Ca2+/calmodulin-dependent protein kinase II by RNA interference.
Hippocampal alpha-Ca2+/calmodulin-dependent protein kinase II (alpha-CaMKII) has been implicated in spatial learning, neuronal plasticity, epilepsy, and cerebral ischemia. In the present study, an adeno-associated virus (AAV) vector was designed to express green fluorescent protein (GFP) from the CBA promoter and a small hairpin RNA targeting alpha-CaMKII (AAV-shCAM) driven from the U6 promoter. The AAV-shCAM or control vector was microinfused into the rat hippocampus and behavioral testing conducted 19-26 days following surgery. Expression of the marker gene and alpha-CaMKII was evaluated 31 days following AAV infusion. GFP expression was localized to the hippocampus and extended +/-2 mm rostral and caudal from the injection site. Hippocampal alpha-CaMKII was significantly reduced following AAV-shCAM treatment as demonstrated using immunohistochemical and Western analysis. This suppression of alpha-CaMKII was associated with changes in exploratory behavior (open field task) and impaired place learning (water maze task). These results demonstrate the efficacy of a viral-based delivered shRNA to produce gene suppression in a specific circuit of the brain.
Babcock
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AM
Department of Psychology, Montana State University, Bozeman, MT 59717, USA. mbabeock@montana.edu
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D000818:Animals; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000032:analysis / Q000037:antagonists & inhibitors* / Q000235:genetics; D000229:Dependovirus / Q000235:genetics*; D005260:Female; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000235:genetics; D006624:Hippocampus / Q000201:enzymology*; D018782:Maze Learning; D011401:Promoter Regions, Genetic / Q000235:genetics; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D012342:RNA, Small Nuclear / Q000235:genetics; D051381:Rats
D034741:RNA, Small Interfering; D012342:RNA, Small Nuclear; C405486:U6 small nuclear RNA; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C517675:Camk2a protein, rat