Figure 1. Glycine inhibition in the dorsal horn is lost in chronic pain states, but may be restored by GlyT2 inhibitors to alleviate pain: Glycinergic inhibition to lamina II excitatory interneurons is lost in chronic pain states, leading to enhancement of nociceptive signals, and aberrant activation of the nociceptive pathways in response to non-noxious stimuli such as touch (via Aβ fibres). Drugs that rectify this glycinergic dysfunction, such as GlyT2-inhibitors, are able to alleviate pain by inhibiting the abnormal signalling.
Figure 2. Analgesia (von Frey) testing in a mouse model of chronic neuropathic (CCI) pain: A Ol-D-lys (purple) produced analgesia in mice in a dose-dependent manner from 10 mg kg-1 to 30 mg kg-1 at which a peak effect was seen. Higher doses of 50 mg kg-1 and 100 mg kg-1 were less effective than the lower dose of 30 mg kg-1. B ORG25543 produced statistically significant analgesia from 60 min to 4 h at the 30 mg kg-1 dose. At the highest tested dose of 50 mg kg-1, responses to the von Frey stimuli were markedly reduced, however this coincided with severe side effects and toxicity, which precluded testing the full data set and this dose was excluded from statistical analysis. Significant post-hoc multiple comparisons are indicated by asterisks (* p < 0.05). n’s per group are indicated in parentheses in legend.
Figure 3. Analgesia (von Frey) testing in a mouse model of inflammatory (CFA) pain: No analgesia was observed for any dose of ol-D-lys or for 30 mg kg-1 of ORG25543. n’s per group are indicated in parentheses in legend.
Figure 4. Acute pain (hotplate) testing in mice: No anti-nociception was observed for 30 mg kg-1 of ol-D-lys or for 30 mg kg-1 of ORG25543 while non-parametric Kruskal-Wallis one-way test showed significant anti-nociception at 30-min in the morphine (orange, 10 mg kg-1) group (* p < 0.05). n’s per group are indicated in parentheses in legend.
Figure 5. Side effects and convulsions caused by GlyT2-inhibitors: A(i)-(ii) show side effects of ol-D-lys. B(i)-(ii) show side effects of ORG25543. C shows convulsions caused by ol-D-lys. D (i)-(ii) convulsions caused by ORG25543. Summary shows mean +/- SEM. Panel (ii) of A, B and D show the scatter of the doses that caused significant adverse behaviours compared to t=0, for which Wilcoxon signed rank test was performed (* p < 0.05). The cross marked after 1 hour in B(i) and D(i) indicates the time at which one animal died. n’s per group are indicated in parentheses in legend.
Figure 6. Motor coordination Rotarod test: AOl-D-lys caused no dose-dependent motor incoordination. However, an enhanced performance was observed at 60 and 90 minutes, at the 16 mg kg-1 group performed (* p < 0.05).B ORG25543 caused profound and immediate incoordination in 2 out of 3 animals at 50 mg kg-1, however due to the low numbers tested, this could not be statistically analysed. n’s per group are indicated in parentheses in legend.
Figure 7. Respiratory side effects: Whole body plethysmography of freely moving mice showed that a sub-maximal analgesic dose of morphine (orange) caused respiratory depression, measured by respiratory frequency and minute volume (MVb), but ol-D-lys did not. Significant post-hoc multiple comparisons are indicated by asterisks (*p < 0.05). n’s per group are indicated in parentheses in legend.