Effects of microinjection of antisense nNOS oligonucleotide into the nucleus tractus solitarii on circulation

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We examined the cardiovascular effects of bilateral microinjection of antisense oligodeoxynucleotides (AS-ODNs) to neuronal nitric oxide synthase (nNOS) into the nucleus tractus solitalii (NTS) of Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR), and compared the results of SHR with those of WKY to investigate the mechanisms of hypertension of SHR. The AS-ODN 5'-CTTCCATGGTATCTGTG-3' was designed to target an initiation codon of rat nNOS mRNA ; the complementary sense sequence was 5'-CACAGATACCATGGAAG-3' ; the scrambled sequence with the same GC content was 5'-CGTAGTAGGCATACGTA-3'. The ODNs were diluted in artificial cerebrospinal fluid and mixed with cationic lipids for transfection. In urethane-anesthetized, paralyzed WKY, bilateral microinjection of nNOS AS-ODNs (20 pmole in a 50 nl volume) into the NTS produced a significant increase in mean arterial blood pressure (MABP) 30-60 min after injection as compared with rats injected with nNOS sense or scrambled ODNs. Similarly, nNOS AS-ODNs produced a significant increase in heart rate 30 min after microinjection compared with nNOS sense ODNs. In SHR, bilateral microinjection of nNOS AS-ODNs into the NTS produced a significant increase in MABP 15 min after injection compared with rats injected with nNOS sense or scrambled ODNs. The pressor responses in SHR were produced earlier than those in WKY. Immunohistochemical study demonstrated that nNOS-immunoreactivity in the rat NTS was suppressed by nNOS AS-ODNs. These results suggest that expression of the nNOS gene in the NTS of WKY and SHR regulates blood pressure.