The three isoforms of nitric oxide synthase distinctively affect mouse nocifensive behavior

Abstract

Nitric oxide synthases (NOSs) have been shown to modulate thermal hyperalgesia and mechanical hypersensitivity in inflammatory and neuropathic pain. However, little is known about the effect of NOSs on baseline function of sensory nerve fibers. Using genetic deficiency and pharmacologic inhibition of NOSs, we examined the impact of the three isoforms NOS1, NOS2, and NOS3 on baseline nocifensive behavior by measuring current vocalization threshold in response to electrical stimulation at 5, 250, 2000 Hz that preferentially stimulate C, Aδ, and Aβ fibers. In response to 5, 250 and 2000 Hz, NOS1-deficient animals had significantly higher current vocalization thresholds compared with wild-type. Genetic deficiency of NOS2 was associated with higher current vocalization thresholds in response to 5 Hz (C-fiber) stimulation. In contrast, NOS3-deficient animals had an overall weak trend toward lower current vocalization thresholds at 5 Hz and significantly lower current vocalization threshold compared with wild-type animals at 250 and 2000 Hz. Therefore, NOSs distinctively affect baseline mouse current vocalization threshold and appear to play a role on nocifensive response to electrical stimulation of sensory nerve fibers.

Fig. 1

Effect of NOS1 deficiency on current vocalization threshold in female (L panels) and male (R panels) mice in response to electrical stimulations at 5, 250 and 2000 Hz that preferentially stimulate C, Aδ, and Aβ sensory fibers respectively. Least square means ± SEM threshold are shown.*indicates 0.01 < p < 0.05, † 0.005 < p ≤ 0.01, and ‡ p ≤ 0.005.

Fig. 2

Effect of NOS2 (L panels) and NOS3 (R panels) on current vocalization threshold in female mice in response to electrical stimulations at 5, 250 and 2000 Hz that preferentially stimulate C, Aδ, and Aβ sensory fibers respectively. Least square means ± SEM threshold are shown.*indicates 0.01 < p < 0.05, † 0.005 < p ≤ 0.01, and ‡ p ≤ 0.005.

Fig. 3

Effect of NOS1 deficiency on NOS2, NOS3, and chemokines gene expression in cerebral cortex and spinal cord. The box plots represent the 25th to 75th interquartile expression ratio (the middle 50% of observations) and the line in the box, the median gene expression ratio comparing NOS1 deficient vs. wild-type animals used as calibrators (expression of 1). Whiskers represent minimum and maximum observations. Numbers below each listed gene represent the p value of expression ratio comparing NOS1-KO vs. wild-type animals. In cerebral cortex, NOS1 deficient animals have significant upregulation of NOS2, NOS3, C–C chemokine receptor type 1 (CCR1), C–C chemokine ligand 3 (CCL3) but similar expressions of C–C chemokine receptor 5 (CCR5) and C–C chemokine ligand 5 (CCL5) compared with wild-type controls. In contrast in spinal cord, NOS1 deficient animals have significant downregulation of NOS2, NOS3, CCL3, CCR5, and CCL5 but similar expression of CCR1 compared with wild-type controls. N = 5 animals per group.