Figure 3: Systemic delivery of GCaMP sensor and excitatory DREADD actuator using AAV-MaCPNS1 enable functional characterization of DRG neurons in mouse models.

A. Illustration of IV administration of MaCPNS1 capsid packaged with ssAAV:CAG-jGCaMP8s genome in mice (~8 week-old young adults, C57BL/6J males, 1×10¹² vg IV dose/mouse, n=4). Three weeks post-expression, the mice were anesthetized and subjected to in vivo calcium imaging in nodose ganglia (NG, top zoom-in), and glucose infusion and distension in the gut (bottom zoom-in). B. Representative images of MaCPNS1 vector-mediated expression of jGCaMP8s (green) in NG in vivo (left), and in post-hoc fixed tissue (right) (scale bar: 100 μm). C. Single-cell activity response measured by calcium signal dynamics in the NG (data pooled from 4 experimental mice). Left panel: NG neuronal response to glucose infusion (white dotted line). Right panel: NG neuronal response to gut distension (white dotted line). D. Illustration of the pain induction experimental workflow. MaCPNS1 vector with ssAAV:hSyn-DIO-hM3D(Gq)-mCherry was intraperitoneally administered to a TrpV1-Cre mouse model (postnatal stage 1 (P1), males, 3×10¹¹ vg IV dose/mouse). After six weeks of expression, the mice (AAV injected or untreated) were habituated and subjected to intraplantar injection with the agonist CNO, after which the mice were monitored for nocifensive lifting/licking behaviors. E. Representative images of DRG sections showing MaCPNS1 vector-mediated mCherry (red) expression. The tissues were co-stained with αNF200 (cyan), αTuj1 (blue) and αCGRP (yellow) markers. F. Total bouts of, and G total time spent lifting or licking the footpad within 15 minutes of injection in uninfected (n=6) and MaCPNS1-infected (n=10) mice treated with CNO. **P ≤ 0.01 by unpaired t-test. Mean+/− sem are shown.