Fig. 1.

In spinal cord of male rats, opioid withdrawal augments MOR-coupled facilitation of evoked EM2 release and MOR coupling to G_s. A, sufentanil inhibited EM2 release from naive spinal tissue (Nv). In contrast, in withdrawn spinal tissue, sufentanil enhanced evoked EM2 release (WD) (n = 3–4). %Δ represents the difference of the percentage increment in K⁺-evoked EM2 release, calculated as [(evoked − basal)/basal] × 100, in the absence and presence of sufentanil. B, sufentanil (1–1000 nM) enhances evoked EM2 release from opioid-withdrawn spinal tissues but this modulation was not dose-dependent (n = 3–5). C, intrathecal PTX unmasks the dose dependence of the facilitation by sufentanil (WD+PTX; 1–100 nM; n = 3), which was abolished by pretreatment of intrathecal CTX in addition to PTX (WD+PTX+CTX; n = 3). D, MOR Western analysis of immunoprecipitates obtained from opioid naive (Nv) and withdrawn (WD) spinal cord using anti-G_sα antibodies. Immunoprecipitation was performed in the absence (−) and presence (+) of 1 μM sufentanil. Sufentanil stimulation of the CoIP of MOR with G_sα was abolished by naloxone (1 μM; WD+Nx). The first two lanes and last three lanes were processed and blotted in parallel. E, dose-dependent stimulation by sufentanil of MOR in G_sα immunoprecipitate (lanes 1–5) and its abolishment by intrathecal CTX (lane 6). All lanes were processed and blotted in parallel. F, quantification of the sufentanil-stimulated increment in MOR G_sα CoIP illustrated in E (n = 3). Opioid withdrawal augments facilitatory MOR-coupled modulation of EM2 release, which is causally associated with increased coupling of MOR with G_s. †, p < 0.05 for withdrawal effect; #, p < 0.05 for sufentanil dose effect; *, p < 0.05 for CTX effect.