Localization of AP2α2, TRPV1 and PIEZO2 to the Large Dense Core Vesicles of Human Dorsal Root Ganglion Neurons

Dorsal Root Ganglia (DRG) consist of both peptidergic and non-peptidergic nociceptive neurons. CGRP, an inflammatory neuropeptide, is a classical marker of peptidergic nociceptors and CGRP is stored within the large dense core vesicles (LDCVs) of these neurons. In addition to storing large peptide neurotransmitters, LDCVs might also serve to transport key membrane proteins to the peripheral terminals. This immunohistochemical study investigated the localization of different membrane proteins to the LDCVs of human DRG neurons. Previously validated antibodies against the endocytotic subunit AP22, the heat-activated channel TRPV1 and the mechanosensitive channel PIEZO2 were used in conjunction with an antibody against CGRP on sections of intact human DRG isolated from de-identified human subjects. Immunohistochemical studies were also performed on human synovial tissue to examine peripheral terminals. High magnification confocal microscopy was used to determine the co-localization signal of these membrane proteins with CGRP. We observed a strong co-localization of AP22 with the CGRP containing LDCVs signifying its role in membrane recycling. Moreover, we also observed a strong colocalization of TRPV1 and PIEZO2 with CGRP suggesting that LDCV release controls the trafficking of these channels to the membrane. It is likely that during injury, bulk exocytosis of CGRP will concomitantly increase the surface expression of TRPV1 and PIEZO2 channels enhancing the responsiveness of these neurons to painful stimuli. This model suggests that neurons that co-localize TRPV1 and PIEZO2 to CGRP containing LDCVs are likely silent nociceptors.