Endocannabinoids acting at the cannabinoid type 1 receptor (CB1R) are known to regulate attention, cognition and mood. Previous studies have shown that, in the rat medial prefrontal cortex (mPFC), CB1R agonists increase norepinephrine release, an effect that may be attributed, in part, to CB1Rs localized to noradrenergic axon terminals. The present study was aimed at further characterizing functional interactions between CB1R and adrenergic receptor (AR) systems in the mPFC using in-vitro intracellular electrophysiology and high-resolution neuroanatomical techniques. Whole-cell patch-clamp recordings of layer V/VI cortical pyramidal neurons in rats revealed that both acute and chronic treatment with the synthetic CB1R agonist WIN 55,212-2 blocked elevations in cortical pyramidal cell excitability and increases in input resistance evoked by the α2-adrenergic receptor (α2-AR) agonist clonidine, suggesting a desensitization of α2-ARs. These CB1R–α2-AR interactions were further shown to be both action potential- and gamma-aminobutyric acid-independent. To better define sites of cannabinoid–AR interactions, we localized α2A-ARs in a genetically modified mouse that expressed a hemoagglutinin (HA) tag downstream of the α2A-AR promoter. Light and electron microscopy indicated that HA-α2A-AR was distributed in axon terminals and somatodendritic processes especially in layer V of the mPFC. Triple-labeling immunocytochemistry revealed that α2A-AR and CB1R were localized to processes that contained dopamine-β-hydroxylase, a marker of norepinephrine. Furthermore, HA-α2A-AR was localized to processes that were directly apposed to CB1R. These findings suggest multiple sites of interaction between cortical cannabinoid–adrenergic systems that may contribute to understanding the effect of cannabinoids on executive functions and mood.

The serotonin (5-hydroxytryptamine, 5-HT) system plays an important role in stress-related psychiatric disorders and substance abuse. Previous work has shown that the dorsal raphe nucleus (DR)-5-HT system is inhibited by swim stress via stimulation of GABA synaptic activity by the stress neurohormone corticotropin-releasing factor (CRF). Additionally, the DR 5-HT system is regulated by opioids. The present study tests the hypothesis that the DR 5-HT system regulates stress-induced opioid relapse. In the first experiment, electrophysiological recordings of GABA synaptic activity in 5-HT DR neurons were conducted in brain slices from Sprague-Dawley rats that were exposed to swim stress-induced reinstatement of previously extinguished morphine conditioned place preference (CPP). Behavioral data indicate that swim stress triggers reinstatement of morphine CPP. Electrophysiology data indicate that 5-HT neurons in the morphine-conditioned group exposed to stress had increased amplitude of inhibitory postsynaptic currents (IPSCs), which would indicate greater postsynaptic GABA receptor density and/or sensitivity, compared to saline-controls exposed to stress. In the second experiment, rats were exposed to either morphine or saline CPP and extinction, and then 5-HT DR neurons from both groups were examined for sensitivity to CRF in vitro. CRF induced a greater inward current in 5-HT neurons from morphine-conditioned subjects compared to saline-conditioned subjects. These data indicate that morphine history sensitizes 5-HT DR neurons to the GABAergic inhibitory effects of stress as well as to some of the effects of CRF. These mechanisms may sensitize subjects with a morphine history to the dysphoric effects of stressors and ultimately confer an enhanced vulnerability to stress-induced opioid relapse.

The cannabinoid receptor agonist, WIN 55,212-2, increases extracellular norepinephrine levels in the rat frontal cortex under basal conditions, likely via desensitization of inhibitory α2-adrenergic receptors located on norepinephrine terminals. Here, the effect of WIN 55,212-2 on stress-induced norepinephrine release was assessed in the medial prefrontal cortex (mPFC), in adult male Sprague-Dawley rats using in vivo microdialysis. Systemic administration of WIN 55,212-2 thirty minutes prior to stressor exposure prevented stress-induced cortical norepinephrine release induced by a single exposure to swim when compared to vehicle. To further probe cortical cannabinoid-adrenergic interactions, postsynaptic α2-adrenergic receptor (AR)-mediated responses were assessed in mPFC pyramidal neurons using electrophysiological analysis in an in vitro cortical slice preparation. We confirm prior studies showing that clonidine increases cortical pyramidal cell excitability and that this was unaffected by exposure to acute stress. WIN 55,212-2, via bath application, blocked postsynaptic α2-AR mediated responses in cortical neurons irrespective of exposure to stress. Interestingly, stress exposure prevented the desensitization of α2-AR mediated responses produced by a history of cannabinoid exposure. Together, these data indicate the stress-dependent nature of cannabinoid interactions via both pre- and postsynaptic ARs. In summary, microdialysis data indicate that cannabinoids restrain stress-induced cortical NE efflux. Electrophysiology data indicate that cannabinoids also restrain cortical cell excitability under basal conditions; however, stress interferes with these CB1-α2 AR interactions, potentially contributing to over-activation of pyramidal neurons in mPFC. Overall, cannabinoids are protective of the NE system and cortical excitability but stress can derail this protective effect, potentially contributing to stress-related psychopathology. These data add to the growing evidence of complex, stress-dependent modulation of monoaminergic systems by cannabinoids and support the potential use of cannabinoids in the treatment of stress-induced noradrenergic dysfunction.