Many ion channels are involved in the pathophysiology of pain, with an important pronociceptive role of Cav3.2 voltage-gated calcium channels in rodents. Cav3.2 channels activate at low membrane potential before the threshold of action potentials and therefore are important contributors to cell excitability. While Cav3.2 is unique in modulating pain signals, the two other low-voltage-gated calcium channels, Cav3.1 and Cav3.3, have similar biophysical properties, overlapping expression, and close sub-cellular distribution including within the spinal cord, the first relay of nociceptive information processing. Therefore we hypothesize that, in addition to its electrogenic impact, Cav3.2 is specifically present in some signaling complexes associated with membrane or submembrane partners that could explain its pro nociceptive functions. Documentation of interactors for this channel family is sparse; and the specific Cav3.2-channelosome is largely unknown.

This study aims to identify Cav3.2-associated signaling complexes with the hope of finding specific interactors. To do so, we use Affinity Purification associated with Mass Spectrometry (AP-MS) on tissues from Cav3.2-GFP KI mice using a set of high affinity anti-GFP dual nanobodies reversibly coupled to magnetic beads.

The combined use of genetically tagged channel, high affinity anti-tag trapping, and specific chemical elution enabled the enrichment of lowly expressed Cav3.2 to an extent compatible with MS analysis. The preliminary set of partners obtained validate this discovery pipeline.

 The most co-enriched partners show promising perspectives in the context of pain circuits. We believe that applying this non-biased strategy in models of chronic pain compared to control situations will enable us to pinpoint perturbations of Cav3.2 complexes that could be targeted by future innovative analgesic therapies for chronic pain management. 

Supported by ANR interacT / FHU inovpain / Labex ICST