Functionally differentiating two neuronal nitric oxide synthase isoforms through antisense mapping: evidence for opposing NO actions on morphine analgesia and tolerance.
Several isoforms of neuronal nitric oxide synthase (nNOS) have been identified. Antisense approaches have been developed which can selectively down-regulate nNOS-1, which corresponds to the full-length nNOS originally cloned from the brain, and nNOS-2, a truncated form lacking two exons which is generated by alternative splicing, as demonstrated by decreases in mRNA levels. Antisense treatment also lowers nNOS enzymatic activity. Down-regulation of nNOS-1 prevents the development of morphine tolerance. Whereas morphine analgesia is lost in control and mismatch-treated mice given daily morphine injections for 5 days, mice treated with antisense probes targeting nNOS-1 show no decrease in their morphine sensitivity over the same time period. Conversely, an antisense probe selectively targeting nNOS-2 blocks morphine analgesia, shifting the morphine dose-response curve over 2-fold to the right. Both systems are active at the spinal and the supraspinal levels. An antisense targeting inducible NOS is inactive. Studies with NG-nitro-L-arginine, which does not distinguish among NOS isoforms, indicate that the facilitating nNOS-2 system predominates at the spinal level while the inhibitory nNOS-1 system is the major supraspinal nNOS system. Thus, antisense mapping distinguishes at the functional level two isoforms of nNOS with opposing actions on morphine actions. The ability to selectively down-regulate splice variants opens many areas in the study of nNOS and other proteins.
Kolesnikov
Y A
YA
The Cotzias Laboratory of Neuro-Oncology and Departments of Neurology and Anesthesiology, Memorial Sloan-Kettering Cancer Center, New York, NY 10021, USA.
Pan
Y X
YX
Babey
A M
AM
Jain
S
S
Wilson
R
R
Pasternak
G W
GW
D000698:Analgesia; D000818:Animals; D004361:Drug Tolerance; D007527:Isoenzymes / Q000378:metabolism*; D008297:Male; D051379:Mice; D009020:Morphine / Q000494:pharmacology*; D009474:Neurons / Q000201:enzymology*; D009569:Nitric Oxide / Q000502:physiology*; D019001:Nitric Oxide Synthase / Q000378:metabolism*; D019335:Nitroarginine / Q000494:pharmacology; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*
D007527:Isoenzymes; D016376:Oligonucleotides, Antisense; D019335:Nitroarginine; D009569:Nitric Oxide; D009020:Morphine; D019001:Nitric Oxide Synthase