Pain modulators regulate the dynamics of PKA-RII phosphorylation in subgroups of sensory neurons

Abstract

Knowledge about the molecular structure of protein kinase A (PKA) isoforms is substantial. In contrast, the dynamics of PKA isoform activity in living primary cells has not been investigated in detail. Using a high content screening microscopy approach, we identified the RIIb subunit of PKA-II to be predominantly expressed in a subgroup of sensory neurons. The RIIb-positive subgroup included most neurons expressing nociceptive markers (TRPV1, NaV1.8, CGRP, IB4) and responded to pain-eliciting capsaicin with calcium influx. Isoform-specific PKA reporters showed in sensory-neuronderived F11 cells that the inflammatory mediator PGE 2 specifically activated PKA-II but not PKA-I. Accordingly, pain-sensitizing inflammatory mediators and activators of PKA increased the phosphorylation of RII subunits (pRII) in subgroups of primary sensory neurons. Detailed analyses revealed basal pRII to be regulated by the phosphatase PP2A. Increase of pRII was followed by phosphorylation of CREB in a PKA-dependent manner. Thus, we propose RII phosphorylation to represent an isoform-specific readout for endogenous PKA-II activity in vivo, suggest RIIb as a novel nociceptive subgroup marker, and extend the current model of PKA-II activation by introducing a PP2A-dependent basal state.

Crosstalk from cAMP to ERK1/2 emerges during postnatal maturation of nociceptive neurons and is maintained during aging

“…We complemented this data by analyzing the expression of two kinase isoforms, PKA-RIIβ and CaMKIIα, involved in pain sensitization of nociceptors (Brüggemann et al, 2000;Chen et al, 2010;Isensee et al, 2014a). With the exception of CaMKIIα, all marker proteins were already detectable at birth, but changed their cellular expression pattern during aging ( Figs 2B and 3).…”

“…We have recently shown that regulatory subunit PKA-RIIβ identifies not only some, but the large majority, if not all, different nociceptive subgroups (Isensee et al, 2014a). If this integrative marker of all nociceptive neurons develops before, after, or simultaneously with the establishment of nociceptive subgroups as defined by Nav1.8 or TRPV1 expression has not been analyzed.…”

“…The PKA regulatory subunit RIIβ (also known as PRKAR2B) is especially enriched in all nociceptive subgroups but not in other sensory subtypes, such as neurofilament 200 (NF200; also known as NEFH)-expressing neurons. Subgrouping through PKA-RIIβ expression allowed a much better and integrative identification of all nociceptors than could be achieved by assessing any of the classical markers of nociceptors (Isensee et al, 2014a). The notion that intracellular sensitization signaling might be a differentiating property of nociceptive neurons was corroborated by a separate study.…”

“…Recently, we found for the first time that the expression of components of the intracellular sensitization signaling, indeed, also defines nociceptive neurons (Isensee et al, 2014a). The PKA regulatory subunit RIIβ (also known as PRKAR2B) is especially enriched in all nociceptive subgroups but not in other sensory subtypes, such as neurofilament 200 (NF200; also known as NEFH)-expressing neurons.…”

“…PKA mediates sensitization induced by inflammatory mediators, such as the Gα scoupled GPCR ligands prostaglandin E2 (PGE 2 ), prostacyclin (PGI 2 ), 5-hydroxytryptamine (5-HT; also known as serotonin) and epinephrine (Isensee et al, 2014a). On the other hand, ERK1/2 is central for growth factor-induced sensitization such as that mediated by NGF (Andres et al, 2010).…”