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The AT1-R has been implicated in many cellular and physiological actions of angiotensin II (AII) in the brain. A retrovirus vector (LNSV) containing an AT1B-R antisense sequence (AT1B-AS) (termed LNSV-AT1B-AS) was constructed and used to determine the feasibility of using viral-mediated gene transfer to control AT1-Rs and AII actions in astroglial and neuronal cells in primary cultures from rat brain. Briefly, a 1.26-kb antisense sequence corresponding to nt -132 to +1128 of AT1-R cDNA was cloned into the LNSV vector, the vector was transfected into PA317 cells, and transfected cells were selected in G418. Incubation of brain cells with culture medium containing LNSV-AT1B-AS viral particles showed that AT1B-AS was integrated into the genome and transcribed in brain cells. This was associated with a significant decrease in AT1-Rs and in the AII-stimulated increase of c-fos mRNA, a measure of AT1-R function. These observations show that the AT1B-AS gene can be transferred into astroglial cells in culture by LNSV and that such a transfer inhibits AT1-Rs and the AII stimulation of cellular activities. In addition, the usefulness of this approach to study AII-dependent pathophysiology in primary neuronal cultures from brain, in particular, is established.

proceedings of the national academy of sciences of the united states of america

Retrovirus-mediated transfer of an angiotensin type I receptor (AT1-R) antisense sequence decreases AT1-Rs and angiotensin II action in astroglial and neuronal cells in primary cultures from the brain.