Electrophysiological Effects of Kappa-Opioid Analgesic, RU-1205, Using Machine Learning Methods

This study focuses on RU-1205, a new kappa-opioid agonist exhibiting analgesic effect without causing dysphoric or aversive reactions. It is assumed that the absence of dysphoric or aversive effects can be attributed to functional selectivity or it might be due to an additional mechanism of action that involves blocking the p38 mitogen-activated protein kinase (MAPK).The aim of this study was the experimental identification of the mechanisms of action of RU-1205 associated with inhibition of MAPK p38 and functional selectivity at kappa-opioid receptors.Materials and methods Rats weighing 260-280 g were implanted with chronic cortical and deep electrodes. LFP activity was recorded after intracerebroventricular administration of well-studied reference substances: the selective kappa-opioid agonist U-50488 at a dose of 100 μg; the MAPK p38 blocker SB203580 at a dose of 1 μg; and the investigational compound RU-1205 at 350 μg. The weighted phase lag index (WPLI) was calculated. Subsequently, machine learning techniques were employed to reduce dimensionality and extract connectivity features using the principal component analysis method. Finally, signal classification was conducted using models based on Gaussian processes.By applying the patch-clamp technique in the whole-cell configuration, the spike activity of pyramidal neurons in the basolateral amygdala was studied. The neurons were identified by their accommodation properties. After local perfusion of the test compounds, 3 dose-response curves were obtained for: (1) U-50488 at concentrations ranging from 0.001 to 10 μM; (2) combinations of U-50488 (0.001–10 μM) and RU-1205 (10 μM); and (3) combinations of U-50488 (0.01–10 μM) and RU-1205 (100 μM).Results The developed models were able to classify the compound RU-1205 as a «non-inhibitor» of MAPK p38 with a probability of 0.89. The results obtained were confirmed in patch clamp experiments on acute brain slices, where it was demonstrated that U-50488 statistically significantly increases the spike activity of pyramidal neurons in the basolateral amygdala (p <0.05) and RU-1205 interacts with U-50488, suppressing its effect on the spike activity of neurons.Conclusions The findings suggest that compound RU-1205 displays properties consistent with a functional kappa opioid receptor agonist and does not have a significant effect on MAPK p38. The study demonstrates the possibility of integrating electrophysiological measurements and advanced data analysis methods for a deep understanding of neuronal mechanisms of drug action and underscores the potential for further research in this area.### Competing Interest StatementThe authors have declared no competing interest.* BBB : blood-brain barrier; CNS : central nervous system; ACSF : artificial cerebrospinal fluid; BLA : basolateral amygdala; LFP : local field potential; p38 MAPK : p38 mitogen-activated protein kinase; WPLI : weighted phase lag index.