Antigen retrieval pre-treatment causes a different expression pattern of Cav3.2 in rat and mouse spinal dorsal horn

Abstract

Ca_v3 channels consist of three isoforms, Ca_v3.1 (α1G), Ca_v3.2 (α1H), and Ca_v3.3 (α1I), which produce low-threshold spikes that trigger burst firings in nociceptive neurons of the spinal dorsal horn (SDH) and dorsal root ganglion (DRG). Although Ca_v3.2 plays a crucial role in pathological pain, its distribution in SDH still remains controversial. One study showed that Ca_v3.2 is ubiquitously expressed in neurons, but another study implied that Ca_v3.2 is expressed restricted to astrocytes. To unravel these discrepancies, we used methods of immunohistochemistry either with or without antigen retrieval (AR) pre-treatment to detect Ca_v3 in SDH and DRG from both rats and mice. Moreover, Ca_v3.2 mRNA was detected in mice SDH using in situ hybridization. We found that the expression pattern of Ca_v3.2 but not Ca_v3.1 and Ca_v3.3 in SDH were largely different with or without AR pre-treatment, which showed a neuron-like and an astrocyte-like appearance, respectively. Double staining further demonstrated that Ca_v3.2 was mainly co-stained with the neuronal marker NeuN in the presence of AR but was with glial fibrillary acidic protein (GFAP, marker for astrocytes) in the absence of AR pre-treatment. Importantly, Ca_v3.2 mRNA was mainly co-localized with Ca_v3.2 but not GFAP. Together, our findings indicate that AR pre-treatment or not impacts the expression pattern of Ca_v3.2, which may make a significant contribution to the future study of Ca_v3.2 in SDH.