Antinociceptive action of oxytocin involves inhibition of potassium channel currents in lamina II neurons of the rat spinal cord

Abstract

Background: Growing evidence in the literature shows that oxytocin (OT) has a strong spinal anti-nociceptive action. Oxytocinergic axons originating from a subpopulation of paraventricular hypothalamic neurons establish synaptic contacts with lamina II interneurons but little is known about the functional role of OT with respect to neuronal firing and excitability.

Results: Using the patch-clamp technique, we have recorded lamina II interneurons in acute transverse lumbar spinal cord slices of rats (15 to 30 days old) and analyzed the OT effects on action potential firing ability. In the current clamp mode, we found that bath application of a selective OT-receptor agonist (TGOT) reduced firing in the majority of lamina II interneurons exhibiting a bursting firing profile, but never in those exhibiting a single spike discharge upon depolarization. Interestingly, OT-induced reduction in spike frequency and increase of firing threshold were often observed, leading to a conversion of the firing profile from repetitive and delayed profiles into phasic ones and sometimes further into single spike profile. The observed effects following OT-receptor activation were completely abolished when the OT-receptor agonist was co-applied with a selective OT-receptor antagonist. In current and voltage clamp modes, we show that these changes in firing are strongly controlled by voltage-gated potassium currents. More precisely, transient IA currents and delayed-rectifier currents were reduced in amplitude and transient IA current was predominantly inactivated after OT bath application.

Conclusion: This effect of OT on the firing profile of lamina II neurons is in good agreement with the antinociceptive and analgesic properties of OT described in vivo.

Figure 1

Firing profiles of lamina II neurons recorded in rat spinal cord slices. A: In the current clamp configuration of the whole-cell patch clamp, four distinct firing profiles were recorded in lamina II neurons in response to incremental depolarizing currents steps of 900 ms every 20 s. They are referred to as repetitive (R), phasic (P), delayed (D) and single spike (S). Typical repetitive firing profile is characterized by the tonic generation of action potentials during the depolarization. The phasic profile is characterized by a strong accommodation leading to a plateau. The delayed firing profile exhibits a time-dependent silent phase before bursting. Lastly, a small proportion of lamina II neurons displays only a single spike (sometimes two) at the beginning of the depolarization. Stars show characteristic firing profile at longer time-scale. Dashed lines represent -60 mV. As shown on the right diagram, no correlation could be found between the firing profile and the location of the recorded neuron. B: Biocytin revelation (images) and reconstruction (drawings) of recorded cells illustrating the large variety of morphology of lamina II neurons. In our hands, no correlation was observed between a specific morphology and the firing pattern.

Figure 2

Oxytocin reduces the firing ability of lamina II neurons. A: Effects of TGOT (1 μM), a selective agonist of OT-receptor bath-applied for 15 minutes, on the different firing profiles expressed by lamina II neurons. Four different examples of firing profile conversion induced by TGOT. Diagram in the grey box gives the scheme of profile conversion observed after TGOT application. B: Example of a lamina II neuron exhibiting a repetitive profile that was not converted after a 15-min application of TGOT. Dashed lines represent -60 mV.

Figure 3

Transient potassium current shapes the firing profile. A: Modulation of the firing profile following the application of 4-AP, a blocker of transient K⁺ currents (I_A), in three different lamina II neurons. B: Firing profile changes exhibited by a phasic neuron (left panels, standard protocol) submitted to conditioning hyperpolarizing (middle panels) or depolarizing prepulse (right panels) aimed at maximizing or minimizing I_A current, respectively. In these three conditions, the effect of TGOT application (bottom traces) is compared to the control situation (upper trace). Black arrowheads indicate the accommodation in spike frequency. White arrowheads show the delay time before obtaining the first action potential. C: Effect of 4-AP on a neuron exhibiting a delayed firing profile after a conditioning hyperpolarizing prepulse producing a large I_A current. Dashed lines represent -60 mV.

Figure 4

Modulation induced by TGOT on properties of sustained (IKDR) and transient (I_A) potassium currents. A: Voltage/current relationship of the sustained (left graph) and transient K⁺ currents (right graph) observed in a lamina II neuron in control (black circles) and after bath application of TGOT (1 μM, white circles). A typical current, obtained at a membrane potential of +50 mV, is shown in inset for both current components in control condition (black line) and after TGOT (dotted line). B, C: Activation (B) and inactivation curves (C) of sustained (left graphs) and transient components of K⁺ currents (right graphs), respectively. Mean data are shown for 11 neurons, each recorded in the control situation (black circles and lines) and after application of 1 μM TGOT (white circles and dotted lines). All experiments were conducted in the presence of a steady-state concentration of TTX (0.5 μM). Values are expressed as means ± S.E.M.