Pharmacological characterisation of the highly NaV1.7 selective spider venom peptide Pn3a

Human genetic studies have implicated the voltage-gated sodium channel NaV1.7 as a therapeutic target for the treatment of pain. A novel peptide, μ-theraphotoxin-Pn3a, isolated from venom of the tarantula Pamphobeteus nigricolor, potently inhibits NaV1.7 (IC50 0.9 nM) with at least 40-1000-fold selectivity over all other NaV subtypes. Despite on-target activity in small-diameter dorsal root ganglia, spinal slices, and in a mouse model of pain induced by NaV1.7 activation, Pn3a alone displayed no analgesic activity in formalin-, carrageenan- or FCA-induced pain in rodents when administered systemically. A broad lack of analgesic activity was also found for the selective NaV1.7 inhibitors PF-04856264 and phlotoxin 1. However, when administered with subtherapeutic doses of opioids or the enkephalinase inhibitor thiorphan, these subtype-selective NaV1.7 inhibitors produced profound analgesia. Our results suggest that in these inflammatory models, acute administration of peripherally restricted NaV1.7 inhibitors can only produce analgesia when administered in combination with an opioid

SLO-2 Is Cytoprotective and Contributes to Mitochondrial Potassium Transport

Mitochondrial potassium channels are important mediators of cell protection against stress. The mitochondrial large-conductance “big” K+ channel (mBK) mediates the evolutionarily-conserved process of anesthetic preconditioning (APC), wherein exposure to volatile anesthetics initiates protection against ischemic injury. Despite the role of the mBK in cardioprotection, the molecular identity of the channel remains unknown. We investigated the attributes of the mBK using C. elegans and mouse genetic models coupled with measurements of mitochondrial K+ transport and APC. The canonical Ca2+-activated BK (or “maxi-K”) channel SLO1 was dispensable for both mitochondrial K+ transport and APC in both organisms. Instead, we found that the related but physiologically-distinct K+ channel SLO2 was required, and that SLO2-dependent mitochondrial K+ transport was triggered directly by volatile anesthetics. In addition, a SLO2 channel activator mimicked the protective effects of volatile anesthetics. These findings suggest that SLO2 contributes to protection from hypoxic injury by increasing the permeability of the mitochondrial inner membrane to K+

Identification of Protein Networks Involved in the Disease Course of Experimental Autoimmune Encephalomyelitis, an Animal Model of Multiple Sclerosis

A more detailed insight into disease mechanisms of multiple sclerosis (MS) is crucial for the development of new and more effective therapies. MS is a chronic inflammatory autoimmune disease of the central nervous system. The aim of this study is to identify novel disease associated proteins involved in the development of inflammatory brain lesions, to help unravel underlying disease processes. Brainstem proteins were obtained from rats with MBP induced acute experimental autoimmune encephalomyelitis (EAE), a well characterized disease model of MS. Samples were collected at different time points: just before onset of symptoms, at the top of the disease and following recovery. To analyze changes in the brainstem proteome during the disease course, a quantitative proteomics study was performed using two-dimensional difference in-gel electrophoresis (2D-DIGE) followed by mass spectrometry. We identified 75 unique proteins in 92 spots with a significant abundance difference between the experimental groups. To find disease-related networks, these regulated proteins were mapped to existing biological networks by Ingenuity Pathway Analysis (IPA). The analysis revealed that 70% of these proteins have been described to take part in neurological disease. Furthermore, some focus networks were created by IPA. These networks suggest an integrated regulation of the identified proteins with the addition of some putative regulators. Post-synaptic density protein 95 (DLG4), a key player in neuronal signalling and calcium-activated potassium channel alpha 1 (KCNMA1), involved in neurotransmitter release, are 2 putative regulators connecting 64% of the identified proteins. Functional blocking of the KCNMA1 in macrophages was able to alter myelin phagocytosis, a disease mechanism highly involved in EAE and MS pathology. Quantitative analysis of differentially expressed brainstem proteins in an animal model of MS is a first step to identify disease-associated proteins and networks that warrant further research to study their actual contribution to disease pathology

Activity of novel lipid glycine transporter inhibitors on synaptic signalling in the dorsal horn of the spinal cord

© 2018 The British Pharmacological Society Background and Purpose: Inhibitory neurotransmission plays an important role in controlling excitability within nociceptive circuits of the spinal cord dorsal horn. Loss of inhibitory signalling is thought to contribute to the development of pathological pain. Preclinical studies suggest that increasing inhibitory glycinergic signalling is a good therapeutic strategy for treating pain. One approach to increase synaptic glycine is to inhibit the activity of the glycine transporter 2 (GlyT2) on inhibitory nerve terminals. These transporters are involved in regulating glycine concentrations and recycling glycine into presynaptic terminals. Inhibiting activity of GlyT2 increases synaptic glycine, which decreases excitability in nociceptive circuits and provides analgesia in neuropathic and inflammatory pain models. Experimental Approach: We investigated the effects of reversible and irreversible GlyT2 inhibitors on inhibitory glycinergic and NMDA receptor-mediated excitatory neurotransmission in the rat dorsal horn. The effect of these drugs on synaptic signalling was determined using patch-clamp electrophysiology techniques to measure glycine- and NMDA-mediated postsynaptic currents in spinal cord slices in vitro. Key Results: We compared activity of four compounds that increase glycinergic tone with a corresponding increase in evoked glycinergic postsynaptic currents. These compounds did not deplete synaptic glycine release over time. Interestingly, none of these compounds increased glycine-mediated excitatory signalling through NMDA receptors. The results suggest that these compounds preferentially inhibit GlyT2 over G1yT1 with no potentiation of the glycine receptor and without inducing spillover from inhibitory to excitatory synapses. Conclusions and Implications: GlyT2 inhibitors increase inhibitory neurotransmission in the dorsal horn and have potential as pain therapeutics. Linked Articles: This article is part of a themed section on Recent Advances in Targeting Ion Channels to Treat Chronic Pain. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.12/issuetoc