Peripheral oxytocin restores light touch and nociceptor sensory afferents towards normal after nerve injury

Abstract

Oxytocin reduces primary sensory afferent excitability and produces analgesia in part through a peripheral mechanism, yet its actions on physiologically characterized, mechanically sensitive afferents in normal and neuropathic conditions are unknown. We recorded intracellularly from L4 dorsal root ganglion neurons characterized as low-threshold mechanoreceptors (LTMRs) or high-threshold mechanoreceptors (HTMRs) in female rats 1 week after L5 partial spinal nerve injury or sham control (n = 24 rats/group) before, during, and after ganglionic perfusion with oxytocin, 1 nM. Nerve injury desensitized and hyperpolarized LTMRs (membrane potential [Em] was -63 ± 1.8 mV in sham vs -76 ± 1.4 mV in nerve injury; P < 0.001), and sensitized HTMRs without affecting Em. In nerve-injured rats, oxytocin depolarized LTMRs towards normal (Em = -69 ± 1.9 mV) and, in 6 of 21 neurons, resulted in spontaneous action potentials. By contrast, oxytocin hyperpolarized HTMRs (Em = -68 ± 2.7 mV before vs -80 ± 3.2 mV during oxytocin exposure; P < 0.01). These effects were reversed after removal of oxytocin, and oxytocin had minimal effects in neurons from sham surgery animals. Sensory afferent neurons immunopositive for the vasopressin 1a receptor were larger (34 ± 6.3 μm, range 16-57 μm) than immunonegative neurons (26 ± 3.4 μm, range 15-43 μm; P < 0.005). These data replicate findings that neuropathic injury desensitizes LTMRs while sensitizing HTMRs and show rapid and divergent oxytocin effects on these afferent subtypes towards normal, potentially rebalancing input to the central nervous system. Vasopressin 1a receptors are present on medium to large diameter afferent neurons and could represent oxytocin's target.

Figure 1.. Experimental method and design

A. Schematic diagram of the L5 partial spinal nerve ligation (pSNL), intracellular recording from neurons in the L4 dorsal root ganglion (DRG) and location of the center of the neuronal receptive fields of the studied afferents in hairy skin of the hindlimb and, lower left, of the glabrous skin of the paw. Circles represent low threshold mechanoreceptors (LTMRs) and triangles represent high threshold mechanoreceptors (HTMRs), with sham represented in black or gray and pSNL in red. Insert at lower right depicts intracellular recording and in vivo perfusion of the DRG with artificial cerebrospinal fluid (aCSF) with and without oxytocin (Oxt) B. Diagram of the activation/stimulation protocol used in this study (RF: receptor field, Em: membrane potential, AP: action potential, Ic: injected current pulses, CV: conduction velocity, MT: Mechanical stimulus above threshold). C. Flowchart and classification of the neurons included in the study and the effects of 5 min exposure to oxytocin (1 nM) on the cellular electrical properties (passive and active) (RA: rapidly adapting, MC: mechanocold).

Figure 2.. Effect of injury and oxytocin on physiologically characterized afferents

A. Effect of artificial cerebrospinal fluid (aCSF) with and without oxytocin (Oxt, 1 nM) of normal rats (black symbols) and those after partial spinal nerve ligation (pSNL; red symbols) and on the somatic membrane potential (Em) of low threshold mechanoreceptor (LTMR; circles) and high threshold mechanoreceptor (HTMR; triangles) afferents. Values are mean ± SE of 12–14 HTMRs and 16–21 LTMRS *=p<0.05; **=p<0.01; ***=p<0.001. B. Representative cellular electrical effects of oxytocin perfusion on LTMRs and HTMRs. Note that the oxytocin exposure induces differential effects in these afferents. In the LTMR (black trace) oxytocin perfusion results in membrane depolarization followed by spontaneous electrotonic potentials (sAPe) and cellular action potentials (sAP) whereas in the HTMR (gray trace) oxytocin perfusion results in membrane hyperpolarization. Lower bars: mechanical stimulation. Calibration bar: 2 ms, 30 mV C. Peripherally evoked somatic action potentials in LTMRs and HTMRs receptors of sham (black traces) and pSNL (red traces) animals and the effects of Oxt (1 nM) exposure on these electrical signatures (gray traces). Calibration bar: 40 mV, 2 sec.

Figure 3.. Pattern of recovery from pain in the study population

A. Representative vasopressin 1A receptor immunoreactivity in sections from L4 dorsal root ganglia (DRGs) visualized by fluorescence microscopy (vasopressin 1a receptors in green and NeuN in red). Calibration bar: 50 μm. B. Greater magnification of the white square area from A. C. Left Panels: Histograms demonstrating distribution of cell size of dorsal root ganglion neurons immunopositive in the entire study population for the vasopressin 1a receptor (V1aR) in black on top and immunonegative neurons in white at bottom. Right Panels: Histograms demonstrating distribution of cell sizes dependent on study group with partial spinal nerve ligation (pSNL) groups in red and sham surgery in black. For each of these groups V1aR immunopositive cells are depicted with filled bars and immunonegative cells with hollow bars.