Protein kinase C in pain: involvement of multiple isoforms

Abstract

Pain is the primary reason that people seek medical care. At present, chronic unremitting pain is the third greatest health problem after heart disease and cancer. Chronic pain is an economic burden in lost wages, lost productivity, medical expenses, legal fees and compensation. Chronic pain is defined as a pain of greater than 2 months duration. It can be of inflammatory or neuropathic origin that can arise following nerve injury or in the absence of any apparent injury. Chronic pain is characterized by an altered pain perception that includes allodynia (a response to a normally non-noxious stimuli) and hyperalgesia (an exaggerated response to a normally noxious stimuli). This type of pain is often insensitive to the traditional analgesics or surgical intervention. The study of the cellular and molecular mechanisms that contribute to chronic pain are of the up-most importance for the development of a new generation of analgesic agents. Protein kinase C isozymes are under investigation as potential therapeutics for the treatment of chronic pain conditions. The anatomical localization of protein kinase C isozymes in both peripheral and central nervous system sites that process pain have made them the topic of basic science research for close to two decades. This review will outline the research to date on the involvement of protein kinase C in pain and analgesia. In addition, this review will try to synthesize these works to begin to develop a comprehensive mechanistic understanding of how protein kinase C may function as a master regulator of the peripheral and central sensitization that underlies many chronic pain conditions.

Figure 1. The pain pathway and its descending modulation

A noxious stimulus will excite peripheral nociceptors (Aδ and C fibers). These fibers synapse on second order dorsal horn neurons. Some of these dorsal horn neurons are excitatory or inhibitory interneurons (in). Others are ascending spinothalamic projection neurons (PN) that ascend via the contralateral ventrolateral funiculus to convey pain sensation to the brain. Dorsal horn neurons are also subject to descending modulation from the midbrain periaquaductal gray (PAG) through polysynaptic circuits through the medulla including through the rostral ventromedial medulla (RVM).

Figure 2. Protein kinase C ε modulates formalin-induced nociception

Subcutaneous administration of dilute 1% formalin in the plantar hindpaw of a one week old rat produces spontaneous flinching and guarding of the hind paw that can be scored in 6 minute bins to provide a Pain Score. A score of 0 represents the absence of paw flinching and a score of 3 represents the presence of flinching or guarding of the hind paw. In panel A, intrathecal administration of an isozyme specific PKCε peptide inhibitor linked to a Tat carrier peptide reduced the average pain score. Attenuation by the PKCε peptide inhibitor could be reversed by co-adminstration with a PKCε peptide activator. The Tat carrier peptide did not alter behaviors as compared to saline or no treatment (148). PKCε peptide activator alone did not increase behaviors with the recognition that behaviors were already maximally stimulated with 1% formalin. In panel B, intrathecal administration of the PKCε peptide activator potentiated pain-associated behaviors stimulated with a sub-maximal concentration of formalin (0.5%). These findings indicate a role for PKCε in nociception.

Figure 3. Protein kinase C ε and γ immunoreactivity in the spinal cord after topical capsaicin

Topical capsaicin was applied to the rat hind paw resulting in thermal hyperalgesia that lasted at least one hour. At the end of the hour rats underwent transcardiac perfusion and lumbar spinal cords were collected for immunohistochemical analysis of protein kinase C ε and γ. In panel A, PKC ε immunoreactivity is seen in the dorsal horn of the spinal cord. The PKC ε appears to be on primary afferent fibers entering into the superficial lamina of the dorsal horn. Using serial spinal cord sections in panel B, PKC γ immunoreactivity is seen in lamina II of the dorsal horn of the spinal cord. In contrast with PKCε, PKCγ is located in dorsal horn neurons.

Figure 4. PKC is positioned to play a central role in peripheral and central sensitization

Panel A, PKC in peripheral sensitization in the primary afferent peripheral terminal. PKC is activated in the primary afferent by a large number of substances that are released in response to injury. These include bradykinin (BK), endothelin-1 (ET), prostaglandins (PGE2), cytokines (tumor necrosis factor, TNF), ATP, trypsin, galanin, and insulin. The receptors for these proteins activate phospholipase C (PLC) and stimulate the production of diacylgycerol (DAG) which activates PKCs. Activated PKC subsequently can increase cation flow into the peripheral terminal via actions on the capsaicin receptor (TRPV1), the acid sensing ion channel (ASIC), and tetrodotoxin-insensitive sodium channels (Nav 1.9). Panel B, PKC is positioned to influence neurotransmitter release from the central primary afferent terminals in the spinal cord. Binding of G-protein coupled pre-synaptic receptors and receptor tyrosine kinases signal through phospholipase C (PLC) to activate PKC. This activated PKC can then enhance calcium influx into the pre-synaptic terminal by phosphorylation of voltage gated calcium channels (VDCC). Simultaneously, PKC can decrease inhibitory tone in the pre-synaptic terminal by phosphorylation of opioid receptors and inhibitory GABA_A receptors effectively decreasing pre-synaptic inhibition. This increase in excitatory tone and decrease in inhibitory tone can potentiate the release of neurotransmitter from the pre-synaptic terminal. Panel C, PKC in the post-synaptic dorsal horn neuron regulates neuronal activity. PKC is activated in dorsal horn neurons subsequent to G protein coupled signaling through phospholipase C (PLC). A variety of G proteins are involved including, but not limited to, neurokinin receptor (NK1), metabotrophic glutamate receptors (mGluR) that bind excitatory amino acids (EAA), and receptors for calcitonin gene-related peptide (CGRP). Activted PKC then phosphorylates Src thus increasing NMDA receptor activity. As in pre-synaptic primary afferent terminals, activated PKC can also modulate post-synaptic opioid receptor activity.

Figure 4. PKC is positioned to play a central role in peripheral and central sensitization

Panel A, PKC in peripheral sensitization in the primary afferent peripheral terminal. PKC is activated in the primary afferent by a large number of substances that are released in response to injury. These include bradykinin (BK), endothelin-1 (ET), prostaglandins (PGE2), cytokines (tumor necrosis factor, TNF), ATP, trypsin, galanin, and insulin. The receptors for these proteins activate phospholipase C (PLC) and stimulate the production of diacylgycerol (DAG) which activates PKCs. Activated PKC subsequently can increase cation flow into the peripheral terminal via actions on the capsaicin receptor (TRPV1), the acid sensing ion channel (ASIC), and tetrodotoxin-insensitive sodium channels (Nav 1.9). Panel B, PKC is positioned to influence neurotransmitter release from the central primary afferent terminals in the spinal cord. Binding of G-protein coupled pre-synaptic receptors and receptor tyrosine kinases signal through phospholipase C (PLC) to activate PKC. This activated PKC can then enhance calcium influx into the pre-synaptic terminal by phosphorylation of voltage gated calcium channels (VDCC). Simultaneously, PKC can decrease inhibitory tone in the pre-synaptic terminal by phosphorylation of opioid receptors and inhibitory GABA_A receptors effectively decreasing pre-synaptic inhibition. This increase in excitatory tone and decrease in inhibitory tone can potentiate the release of neurotransmitter from the pre-synaptic terminal. Panel C, PKC in the post-synaptic dorsal horn neuron regulates neuronal activity. PKC is activated in dorsal horn neurons subsequent to G protein coupled signaling through phospholipase C (PLC). A variety of G proteins are involved including, but not limited to, neurokinin receptor (NK1), metabotrophic glutamate receptors (mGluR) that bind excitatory amino acids (EAA), and receptors for calcitonin gene-related peptide (CGRP). Activted PKC then phosphorylates Src thus increasing NMDA receptor activity. As in pre-synaptic primary afferent terminals, activated PKC can also modulate post-synaptic opioid receptor activity.