HCN1 and HCN2 in rat DRG neurons: levels in nociceptors and non-nociceptors, NT3-dependence and influence of CFA-induced skin inflammation on HCN2 and NT3 expression

CRISTIAN ACOSTA; SIMON MCMULLAN; LAICHE DJOUHRI; LINLIN GAO; ROGER WATKINS; C. BERRY; KATHERINE DEMPSEY; SALLY LAWSON

PLOS ONE

PUBLIC LIBRARY SCIENCE

Lugar: San Francisco; Año: 2012 vol. 7 p. 50442 - 50459

1932-6203

Ih, which influences neuronal excitability, has recently been measured in vivo in sensory neuron subtypes in dorsal root ganglia (DRGs). However, expression levels of HCN (hyperpolarization-activated cyclic nucleotide-gated) channel proteins that underlie Ih were unknown. We therefore examined immunostaining of the most abundant isoforms in DRGs, HCN1 and HCN2 in these neuron subtypes. This immunostaining was cytoplasmic and membrane-associated (ring). Ring-staining for both isoforms was in neurofilament-rich A-fiber neurons, but not in small neurofilament-poor C-fiber neurons, although some C-neurons showed cytoplasmic HCN2 staining. We recorded intracellularly from DRG neurons in vivo, determined their sensory properties (nociceptive or low-threshold-mechanoreceptive, LTM) and conduction velocities (CVs). We then injected fluorescent dye enabling subsequent immunostaining. For each dye-injected neuron, ring- and cytoplasmic-immunointensities were determined relative to maximum ring-immunointensity. Both HCN1- and HCN2-ring-immunointensities were positively correlated with CV in both nociceptors and LTMs; they were high in Abeta-nociceptors and Aalpha/beta-LTMs. High HCN1 and HCN2 levels in Aalpha/beta-neurons may, via Ih, influence normal non-painful (e.g. touch and proprioceptive) sensations as well as nociception and pain. HCN2-, not HCN1-, ring-intensities were higher in muscle spindle afferents (MSAs) than in all other neurons. The previously reported very high Ih in MSAs may relate to their very high HCN2. In normal C-nociceptors, low HCN1 and HCN2 were consistent with their low/undetectable Ih. In some C-LTMs HCN2-intensities were higher than in C-nociceptors. Together, HCN1 and HCN2 expressions reflect previously reported Ih magnitudes and properties in neuronal subgroups, suggesting these isoforms underlie Ih in DRG neurons. Expression of both isoforms was NT3-dependent in cultured DRG neurons. HCN2-immunostaining in small neurons increased 1day after cutaneous inflammation (CFA-induced) and recovered by 4days. This could contribute to acute inflammatory pain. HCN2-immunostaining in large neurons decreased 4days after CFA, when NT3 was decreased in the DRG. Thus HCN2-expression control differs between large and small neurons.