The real time dynamics of vanilloid-induced cytotoxicity and the specific deletion of nociceptive neurons expressing the wild-type vanilloid receptor (VR1) were investigated. VR1 was C-terminally tagged with either the 27-kDa enhanced green fluorescent protein (eGFP) or a 12-amino acid ⑀-epitope. Upon exposure to resiniferatoxin, VR1eGFP-or VR1⑀-expressing cells exhibited pharmacological responses similar to those of cells expressing the untagged VR1. Within seconds of vanilloid exposure, the intracellular free calcium ([Ca 2؉ ] i ) was elevated in cells expressing VR1. A functional pool of VR1 also was localized to the endoplasmic reticulum that, in the absence of extracellular calcium, also was capable of releasing calcium upon agonist treatment. Confocal imaging disclosed that resiniferatoxin treatment induced vesiculation of the mitochondria and the endoplasmic reticulum (ϳ1 min), nuclear membrane disruption (5-10 min), and cell lysis (1-2 h). Nociceptive primary sensory neurons endogenously express VR1, and resiniferatoxin treatment induced a sudden increase in [Ca 2؉ ] i and mitochondrial disruption which was cell-selective, as glia and non-VR1-expressing neurons were unaffected. Early hallmarks of cytotoxicity were followed by specific deletion of VR1-expressing cells. These data demonstrate that vanilloids disrupt vital organelles within the cell body and, if administered to sensory ganglia, may be employed to rapidly and selectively delete nociceptive neurons.

No abstract

Selective blockade of nociceptive pathways represents a mechanism-based approach that has attracted a large variety of pharmacological and molecular investigations. A potential site for selective intervention is the primary afferent nociceptive nerve terminal. Binding of resiniferatoxin (RTX) to the vanilloid-1 receptor (VR1) stimulates and then inactivates heat and vanilloid-responsive nerve endings involved in heat and inflammatory pain signaling which can progress to localized degeneration of the peripheral ending followed by regeneration. Application of RTX directly to peripheral nerve endings produces a long term, reversible attenuation of nociceptive transmission. Heat hyperalgesia and mechanical allodynia were assessed prior to injection of RTX into the hindpaw (baseline) and at acute (minutes-hours) and more chronic (days-weeks) times after injection. Acutely, an inverse dose-to-pain response (guarding, licking) for RTX (0.0625-2.0 microg) occurs, followed by selective attenuation of peripheral pain transmission. Thermal nociception was decreased in a concentration-dependent fashion and lasted up to 21 days, without impairing motor function. Administration of RTX blocked both inflammation-induced hyperalgesia and spinal c-Fos induction. The results demonstrate the efficacy and therapeutic potential of reversible, peripheral C-fiber 'inactivation' for intermediate duration pain control.

The vanilloid receptor type 1 (VR1) is a heat-activated ionophore preferentially expressed in nociceptive neurons of trigeminal and dorsal root ganglia (DRG). VR1, which binds and is activated by capsaicin and other vanilloid compounds, was noted to interact with the endocannabinoid anandamide (ANA) and certain inflammatory metabolites of arachidonic acid in a pH-dependent manner. At pH < 6.5 ANA induced