A Role for Keratinocytes in Peripheral Kappa Opioid Receptor‐Mediated Antinociception
Authors: Pando Miryam M. , Jennings Elaine M. , Chavera Teresa A. , Clarke William P. , Berg Kelly A. 
Published: April 2020

Peripherally‐expressed kappa opioid receptors (KOR) are a promising target to treat inflammatory pain that would be devoid of CNS related adverse effects. We have found that local injectionof KOR agonists into the rat hindpaw produces a profound antinociceptive response that is mediated by activation of G protein‐coupled inwardly‐rectifying potassium (GIRK) channels. We confirmed with immunohistochemistry that both KOR and GIRK2 channels are expressed in intraepidermal nerve fibers (IENFS), which propagate nociceptive signals from the periphery to the spinal cord. Interestingly, we also found that KOR and GIRK2 channels are expressed in keratinocytes. Keratinocytes make up the majority of cells in the epidermis and evidence suggests that they play a role in pain and sensory processing. A variety of nociceptive signaling receptors are expressed in keratinocytes, including transient receptor potential vanilloid channels 1 through 4 (TRPV1‐4), suggesting that keratinocytes can modulate and possibly initiate nociception in the epidermis. Here we sought to determine if expression of GIRK channels in keratinocytes plays a role in KOR‐mediated peripheral antinociception. We assessed the effect of local hindpaw administration of a GIRK2 shRNA expression plasmid fused to a HIV‐1 Tat peptide that facilitates transport into cells. Unlike IENFs, keratinocytes are capable of transcription, thus we used this method to ensure knockdown would occur in keratinocytes and not in IENFs. Rats were injected intraplantarly (i.pl., 50ul) with either GIRK2 shRNA or scrambled non‐effective (sNE) shRNA plasmid complex along with Fugene HD daily for two days. 48 hours after the second injection, the rats were injected i.pl. with ML297 (33ug), a direct GIRK channel activator, and paw withdrawal latencies (PWL) to a thermal stimulus were determined over a 20 min period. ML297 produced a robust (+5 sec above baseline) antinociceptive response in contralateral (non‐shRNA plasmid injected) paws and in hindpaws injected with the sNE shRNA plasmid. Theantinociceptive response to ML297 was lost completely in hindpaws injected with GIRK2 shRNA plasmid. We next determined if local knockdown of GIRK2 in keratinocytes altered the behavioral response to the KOR agonist, U50,488. Following administration of either GIRK2 shRNA or sNE shRNA plasmid, rats were injected with PGE2 (i.pl., 0.3ug) with or without U50,488 (0.1ug) and PWL to the heat stimulus was measured. The sNE shRNA plasmid did not alter the KOR agonist‐mediated reduction of PGE2‐evoked thermal allodynia, however, the GIRK2 shRNA completely blocked the KOR‐mediated response. PGE2‐evoked thermal allodynia in the absence of agonist was unaltered in hindpaws treated with either shRNA plasmid. These data suggest that GIRK channels expressed in keratinocytes play an essential role in peripheral KOR‐mediated antinociceptive signaling in the rat hindpaw.Support or Funding InformationThis work is supported by NIH/NIDA RO1 DA 038645.