Peripheral ablation of type III adenylyl cyclase induces hyperalgesia and eliminates KOR-mediated analgesia in mice
Wen-Wen Zhang, Hong Cao, Yang Li, Xian-Jun Fu, Yu-Qiu Zhang
Wen-Wen Zhang, Hong Cao, Yang Li, Xian-Jun Fu, Yu-Qiu Zhang
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Research Article Neuroscience

Peripheral ablation of type III adenylyl cyclase induces hyperalgesia and eliminates KOR-mediated analgesia in mice

Abstract

Ca2+/calmodulin-stimulated group I adenylyl cyclase (AC) isoforms AC1 and AC8 have been involved in nociceptive processing and morphine responses. However, whether AC3, another member of group I ACs, is involved in nociceptive transmission and regulates opioid receptor signaling remains elusive. Here, we report that conditional KO of AC3 (AC3 CKO) in L3 and L4 DRGs robustly facilitated the mouse nociceptive responses, decreased voltage-gated potassium (Kv) channel currents, and increased neuronal excitability. Furthermore, we report AC3 CKO eliminated the analgesic effect of κ-opioid receptor (KOR) agonist and its inhibition on Kv channel by classical Gαi/o signaling or nonclassical direct interaction of KOR and AC3 proteins. Interestingly, significantly upregulated AC1 level and cAMP concentration were detected in AC3-deficient DRGs. Inhibition of AC1 completely reversed cAMP upregulation, neuronal excitability enhancement, and nociceptive behavioral hypersensitivity in AC3-CKO mice. Our findings suggest a crucial role of peripheral AC3 in nociceptive modulation and KOR opioid analgesia.

Authors

Wen-Wen Zhang, Hong Cao, Yang Li, Xian-Jun Fu, Yu-Qiu Zhang

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Figure 2

Conditional KO of AC3 in L3 and L4 DRGs facilitates nociceptive behavioral responses in mice.

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Conditional KO of AC3 in L3 and L4 DRGs facilitates nociceptive behavior...
(A) Western blot analysis showing a significant decrease in AC3 in L3 and L4 DRGs of CKO mice. The intensity of 3 bands including 180 kDa (glycosylated form), 130 kDa (full-length unglycosylated form), and 70 kDa (monomer form) of AC3 were calculated. *P < 0.05; 2-tailed Student’s t test; n = 5 Ctrl and CKO (mice). (B) AC3-immunoreactivity (AC3-IR, red) colocalized (Co) with EGFP (left) but not with EGFP-T2A-Cre (right) in DRG neurons. Scale bar: 50 μm. (C) Quantitative analysis showing proportion of AC3+ neurons infected with AAV and AC3 knockdown efficiency. (D) Schematic of protocol for virus injection and behavior tests. (E and F) AC3 CKO induced mechanical allodynia in von Frey (E) and paintbrush (F) tests. *P < 0.05, **P < 0.01, 2-tailed Student’s t test; n = 8 Ctrl and CKO (mice). (G) AC3 CKO induced thermal hyperalgesia. **P < 0.01, ****P < 0.0001; 2-way RM ANOVA followed by Bonferroni’s test; n = 7 Ctrl and 11 CKO (mice). (H–K) AC3 CKO facilitated nociceptive responses in 52°C (H) and 55°C (I) hot plate, noxious pinch (J) and intraplantar injection (i.pl.) of 0.1% capsaicin (K). *P < 0.05, **P < 0.01, ***P < 0.001; 2-tailed Student’s t test; n = 9 Ctrl and 10 CKO (mice). (L) I.pl. 2.5% formalin induced a significantly increased nociceptive response at the I phase in AC3-CKO mice. *P < 0.05, ****P < 0.0001; 2-way RM ANOVA followed by Bonferroni’s test (left) and 2-tailed Student’s t test (middle and right); n = 7 Ctrl and CKO (mice). BL, baseline; Ctrl, Control; CKO, AC3 CKO.