Journal: MedComm

Article Title: Both Genetic Manipulation and Acupuncture Alleviate Depressive‐Like Behavior Mediated by Astrocytic P2X7 Receptors

doi: 10.1002/mco2.71042

Figure Lengend Snippet: Treatment with EA prevents the CUMS‐induced increase in expression of P2X7Rs. Determination by quantitative (q)PCR or Western blot of mRNA and protein for P2X7Rs, the purine‐metabolizing enzymes CD39 and CD73, the exocytotic vesicular proteins SNAP23 and SNAP25, and the ATP cellular release protein CALHM2 in mPFC brain slices of mice. p2x7, cd39, cd73, snap23, snap25, calhm2, mRNAs for the respective proteins. Determinations were under control conditions in the medial prefrontal cortex (mPFC) prepared from control mice, as well as in the mPFC of mice treated with CUMS either alone or in combination with EA or sham EA. (A–F) Expression of mRNAs as measured by the ΔΔCt method with respect to β‐actin‐RNA. Relative amounts of p2x7r (A), cd39 (B), cd73 (C), snap23 (D), snap25 (E), and calhm2 (F) mRNAs. (G–L) Representative immunoblots and quantification of the proteins for P2X7R (G), CD39 (H), CD73 (I), SNAP23 (J), SNAP25 (K), and CALHM2 (L) as a ratio of the spot of glycerinaldehyd‐3‐phosphat‐dehydrogenase (GAPDH). (M) Expression of p2x7R mRNA in control mice after EA or sham EA1 (see Section ) treatment. (N) Expression of P2X7 protein in control mice after EA or sham EA1 treatment. All data are shown as mean ± SEM. One‐way ANOVA was followed by the Holm–Sidak test for statistical analysis in case of normal distribution (A–L); * p < 0.05, ** p < 0.01 compared to the control group. The number of individual animals is indicated in each column.

Article Snippet: Membranes were blocked with 5% skimmed milk in Tris‐buffered saline with Tween 20 for 1 h at room temperature and subsequently incubated overnight at 4°C with the following primary antibodies: P2X7R (1:1000; Abcam, Cambridge, UK), CD39 (1:1000; Abcam), CD73 (1:1000; Proteintech, Rosemont, USA), SNAP23 (1:1000; Proteintech), SNAP25 (1:1000; Abcam), CALHM2 (1:400; Proteintech), and GAPDH (1:1000; Proteintech).

Techniques: Expressing, Western Blot, Control, Analysis