Protein Nerve Pain Killer?

Today's post from painresearchforum.org (see link below) is one for the purists and difficult for ordinary people to understand. Nevertheless, if you can plough your way through it you will have a better understanding of some of the directions research into neuropathy is going at the moment. Basically, a new protein has been discovered which can calm down the receptors in the brain and spine which turn your nerves into hyperactive and very excitable pain givers. Researchers have long known that if they could dampen down the excitability of so-called Gaba receptors, they will reduce the typical symptoms of neuropathy considerably. It has been very difficult to do this up to now. They know what to do and where to do it but they don't know how. This protein may well be one of the answers. For more information, you'll need to tackle the article!

Novel Protein Promotes GABAergic Inhibition, Analgesia in Peripheral Nervous System
GINIP regulates GABAB receptor signaling in nociceptors 
by Stephani Sutherland on 26 Sep 2014

With neuropathic pain, the spinal cord suffers a loss of “inhibitory tone”—signaling by GABA, the nervous system’s main inhibitory transmitter. GABA receptors would seem an ideal target to dampen neuropathic hyperexcitability, but considerable efforts toward that aim have largely failed, mainly because GABA receptors are found throughout the nervous system. Now, researchers have identified a novel protein expressed specifically in nociceptors that acts as a key player in analgesic signaling through GABAB receptors. The protein, Gα inhibitory interacting protein (GINIP), could offer a new line of attack to boost GABA transmission by targeting peripheral neurons selectively, possibly avoiding side effects of system-wide GABAB activation.

The work was led by Aziz Moqrich at Centre National de la Recherche Scientifique (CNRS), Paris, France, and was published online September 18 in Neuron.

Ken McCarson, University of Kansas, Kansas City, US, who was not involved in the work, expressed enthusiasm about the team’s identification of GINIP. “There are attributes of GABA signaling that we have been aware of without understanding the mechanisms that underlie them. This work makes an exciting contribution to our understanding of the regulation of endogenous inhibition in the context of persistent pain,” McCarson told PRF.

In the study, co-first authors Stéphane Gaillard and Laure Lo Re set out to identify proteins that define functional subclasses of sensory neurons. Using expression microarrays, they identified several hundred candidates, including GINIP, a protein of unknown function whose expression was restricted to a subset of small, unmyelinated C-fiber neurons. GINIP marked two distinct subgroups of nociceptors: one that expressed isolectin B4 (IB4) and the G protein MRGPRD (Mas-related G protein-coupled receptor member D); these fit the profile of cutaneous free nerve endings. The second group lacked IB4 and expressed markers identifying them as C-fiber low-threshold mechanosensors.

Using a yeast-two-hybrid screen, the researchers discovered that GINIP associated specifically with the G-alpha subunit of inhibitory G protein (Gαi). That finding, Moqrich said, encouraged them to search for a functional role for GINIP in nociceptors.

To do that, the researchers created transgenic mice lacking the protein. “When we knocked out GINIP, the loss did not interfere with temperature or chemical sensation, but we saw a clear phenotype specifically in mechanical pain sensation,” Moqrich told PRF. After either carrageenan injection or nerve injury, all mice developed mechanical hypersensitivity, but in mice lacking GINIP it lasted days or weeks longer than in mice with GINIP.

Because they knew that GINIP interacted with Gαi, the researchers next investigated GABAB receptors, inhibitory G protein-coupled receptors found on nociceptors that are important in pain regulation. In wild-type mice with nerve injury pain, the GABAB receptor agonist baclofen alleviated mechanical pain, but not in GINIP-deficient mice. “Without GINIP, you completely lose the possibility to reverse pain with injection of baclofen,” Moqrich said.

Nociceptors contain two other prominent pain-related receptors coupled to Gαi—the delta and mu opioid receptors (DOR, MOR). In both carrageenan-inflamed and nerve-injured mice, intrathecal injection of DOR or MOR agonists reversed mechanical hypersensitivity in mice whether or not they expressed GINIP. The results suggest a specific role for GINIP acting selectively at GABAB receptors in neuropathic mechanical hypersensitivity independent of opioid receptors.

On nociceptors, GABAB receptors are concentrated at the central terminals, which make synaptic contacts with interneurons in lamina II of the dorsal horn. When those receptors see GABA—or baclofen—G protein signaling results in decreased release of the excitatory neurotransmitter glutamate. In experiments in both isolated sensory neurons and in acute spinal cord slices, the researchers showed that GINIP was required for that inhibitory signaling.



Immunostaining reveals GINIP in a subset of nonpeptidergic nociceptors that targets lamina II in the spinal cord. The left panel shows half of a spinal cord cross-section stained with GINIP antibody (blue), isolectin B4 (IB4, red), and PKCγ (green). The right panel shows a DRG cross-section stained with GINIP antibody (blue), IB4 (red), and TAFA4 (green). The GINIP+IB+ (purple) neurons are the cutaneous free nerve endings, and the GINIP+/TAFA4+ (light blue) neurons are the C-LTMRs. Credit: S. Gaillard, L. Lo Re and A. Moqrich.

How could GINIP facilitate inhibition? When an agonist binds a G protein-coupled receptor, the Gα subunit binds GTP and becomes activated, and the Gβγ subunit is freed up to interact with its downstream targets. Once GTP is cleaved, the complex reassembles and signaling ceases. Importantly, GINIP only bound to the active, GTP-bound form of Gαi and not other types of Gα subunits. Based on their findings, Moqrich postulates that GINIP stabilizes and sequesters the active Gαi subunit, prolonging inhibitory signaling by Gβγ.

Part of what makes GABA receptors so elusive as analgesic targets is the sedative effect of existing receptor agonists that also activate GABA receptors in the brain, explained McCarson. Because GINIP is restricted to nociceptors—the very neurons relevant to neuropathic pain—targeting the GABAB -GINIP interaction offers the possibility of a peripherally restricted therapeutic agent that specifically interferes with GABAB signaling related to pain. “We are at the very embryonic stage of understanding this system,” said Moqrich, but he ventured a guess that such a molecule might take the form of a peptide or a small molecule that mimics or stabilizes binding between GINIP and Gαi. Although the protein interaction is selective to peripheral sensory neurons, it takes place in the cells’ central terminals inside the spinal cord, so it would have to somehow reach those terminals. Despite certain hurdles, the possibility of such an agent offers the hope of dampening pain without touching wakefulness.

“Therapeutics are a long, long way off, but these findings give us something to hang our hats on when it comes to how GABAB signaling is regulated in peripheral nociceptors,” said McCarson.

Stephani Sutherland, PhD, is a neuroscientist, yogi, and freelance writer in Southern California, US.

Image credit: S. Gaillard, L. Lo Re, and A. Moqrich
References:
Editors' Pick
GINIP, a Gαi-Interacting Protein, Functions as a Key Modulator of Peripheral GABAB Receptor-Mediated Analgesia.
Gaillard S, Lo Re L, Mantilleri A, Hepp R, Urien L, Malapert P, Alonso S, Deage M, Kambrun C, Landry M, Low SA, Alloui A, Lambolez B, Scherrer G, Le Feuvre Y, Bourinet E, Moqrich A
Neuron. 2014 Oct 1; 84(1):123-36. Epub 2014 Sep 18.

http://www.painresearchforum.org/news/45895-novel-protein-promotes-gabaergic-inhibition-analgesia-peripheral-nervous-system