INTRODUCTION: Primary nociceptors are the first neurons involved in processing normal and pathological pain. Novel biological approaches such as gene therapies targeting these neurons have great potential, but delivery can be challenging when open or percutaneous infusion into the dorsal root ganglion (DRG) is needed. METHODS: Primary nociceptors were first targeted unilaterally via right hind paw injection of a retrograde adeno-associated virus serotype 2 (AAVRetro) or a mosaic AAVRetro/rh10 vector encoding a constitutive hM3Dq chemogenetic transgene. A second cohort of mice received the same vectors via unilateral intraneural injection in the right sciatic nerve. Changes in pain-related behaviors were assessed 45-60 min following intraperitoneal administration of the ligand clozapine-n-oxide (CNO) using the Von Frey and Hargreaves tests every other week for 6 weeks post-AAVRetro injection. RESULTS: Immunofluorescence and RNA in-situ hybridization of paw injected mice revealed transduction of primary nociceptors in the DRG from L4 to L6 segments, including TrkA, Trpv1, CGRP, and NaV1.8 positive neurons. Chemogenetic activation elicited robust mechanical allodynia responses at 4-week post-RetroAAV delivery in paw injected mice, whilst in sciatic injected mice we observed an earlier pain phenotype starting at 2-week post-RetroAAV administration. No thermal hyperalgesia responses were observed. CONCLUSIONS: We demonstrate for the first time that AAVRetro effectively transduces DRG nociceptors via paw injection. Furthermore, chemogenetic activation caused a strong pain phenotype similar to mice injected in the sciatic nerve. These results support further exploration of non-invasive delivery of retrograde viral vectors to modulate pain neurons for potential therapeutic applications.