Pharmacological studies on the involvement of opioid - nitric oxide (NO) interactions during stress induced anxiogenesis in rats

K Gulati, A Ray. V.P.Chest Institute, University of Delhi, Delhi, Department of Pharmacology, 110007, India

Complex neurochemical pathways in the brain are involved in the regulation of stress induced biological responses. Endogenous opioids are important neuromodulators in the brain and have been implicated in a variety of psychophysiological states. Involvement of various types of opiate receptors (µ, k and δ) and their differential regulatory influence on physiological functions and pathophysiological states are also reported. Interactions of endogenous opioids with other neurotransmitter systems are also reported. Nitric oxide (NO), a unique messenger molecule with multifaceted actions on various organ systems, is also known to act as a crucial neuroregulator in the CNS. Earlier studies have indicated a role for NO during stress and NO modulators have been shown to influence restraint stress induced behavioral, neuroendocrinal and immune responses. The present study evaluated the involvement of opioidergic mechanisms during stress induced anxiogenesis and investigated any possible opioid-NO interactions during such neurobehavioral states in Wistar rats (n=6 per group). Restraint stress (RS) was used as the experimental stressor and anxiety like behavior was assessed by the elevated plus maze (EPM) test. Brain homogenates were then prepared and assayed for stable NO metabolites (NOx) by spectrophotometric method. All drugs were administered intraperitoneally in a volume of 1ml/kg. Single RS (RS x 1) exposure for 1 hour consistently induced an anxiety-like response in the EPM test, i.e. reduced number of open arm entries and time spent in the open arms (Control 23.2±2.9 vs RS 6.9±1.5 for % open arm entries; Control 10.3±0.8 vs RS- 1.1± 0.2 for %open arm time, p<0.05 in each case, ANOVA test) and also elevated the plasma corticosterone levels by 150% as compared to (no RS) controls. Pretreatment with the opioid agonists, morphine (µ), SNC-80 (δ), and to a much lesser extent, U-50488 (κ), attenuated the RS-induced anxiogenic response. Both morphine and SNC-80 increased open arm entries by 111% and 56% respectively as compared to that of corresponding RS group. Such RS induced neurobehavioral suppression was associated with reductions in brain NOx levels (Control 3.0±0.7 vs RS 1.8±0.3), which were also reversed with morphine, and to some extent with SNC-80 treatment. Interaction studies of opioid agonists with NO modulators showed that sub-effective doses of morphine and L-arginine (a NO precursor) had synergistic effects on RS-induced EPM activity, whereas, L-NAME (a NO synthase inhibitor) neutralized the anxiolytic effects of morphine. In another set of experiments, repeated RS (x5) induced adaptative changes - as evidenced by normalization of behavioral suppression in the EPM and elevations in brain NOx, as compared to that seen in RS (x1) group. Pretreatment with morphine, and to a lesser extent SNC 80 or U-50488H, in combination with repeated RS (x5) showed potentiating effects in the induction of behavioral adaptation in the EPM and elevations in brain NOx, as compared to RS (x5) alone. Further, L-NAME, when administered prior to morphine, blocked this effect of the opioid on stress adaptation. These results suggest differential opioid receptor involvement during stress induced modulation of anxiety and that opioid-NO interactions may contribute to such effects.