The effects of polarizing current on nerve terminal impulses recorded from polymodal and cold receptors in the guinea-pig cornea

Abstract

It was reported recently that action potentials actively invade the sensory nerve terminals of corneal polymodal receptors, whereas corneal cold receptor nerve terminals are passively invaded (Brock, J.A., S. Pianova, and C. Belmonte. 2001. J. Physiol. 533:493-501). The present study investigated whether this functional difference between these two types of receptor was due to an absence of voltage-activated Na(+) conductances in cold receptor nerve terminals. To address this question, the study examined the effects of polarizing current on the configuration of nerve terminal impulses recorded extracellularly from single polymodal and cold receptors in guinea-pig cornea isolated in vitro. Polarizing currents were applied through the recording electrode. In both receptor types, hyperpolarizing current (+ve) increased the negative amplitude of nerve terminal impulses. In contrast, depolarizing current (-ve) was without effect on polymodal receptor nerve terminal impulses but increased the positive amplitude of cold receptor nerve terminal impulses. The hyperpolarization-induced increase in the negative amplitude of nerve terminal impulses represents a net increase in inward current. In both types of receptor, this increase in inward current was reduced by local application of low Na(+) solution and blocked by lidocaine (10 mM). In addition, tetrodotoxin (1 microM) slowed but did not reduce the hyperpolarization-induced increase in the negative amplitude of polymodal and cold nerve terminal impulses. The depolarization-induced increase in the positive amplitude of cold receptor nerve terminal impulses represents a net increase in outward current. This change was reduced both by lidocaine (10 mM) and the combined application of tetraethylammomium (20 mM) and 4-aminopyridine (1 mM). The interpretation is that both polymodal and cold receptor nerve terminals possess high densities of tetrodotoxin-resistant Na(+) channels. This finding suggests that in cold receptors, under normal conditions, the Na(+) conductances are rendered inactive because the nerve terminal region is relatively depolarized.

Figure 1.

Effects of polarization on NTIs recorded from cold receptors (A) and polymodal receptors (B). (A and B) The top traces are averaged NTIs recorded in single experiments and the bottom graphs show the mean effects of polarizing current on the positive (fill bars) and negative (open bars) amplitude of NTIs (cold receptors, n = 28; polymodal receptors, n = 17). In the graphs, the positive and negative amplitudes of NTIs were normalized to the positive amplitude measured in the absence of polarizing current (zero current). Statistical comparisons were made with paired sign tests. ***, P < 0.001.

Figure 2.

Effects of varying polarizing current strength on NTI shape in a cold receptor. (A) Overlaid averaged NTIs recorded under control conditions (three traces overlaid) and during application of hyperpolarizing current (30, 40, 60, 80, and 90 nA overlaid) and depolarizing current (−30, −50, −70, −90 nA overlaid). (B) The traces on the left show the pattern of NTI occurrence in the absence of polarizing current and during application of 60 and 90 nA. On the right, averaged NTIs recorded under each condition are shown. During application of 60 nA, the negative amplitude of all NTIs was increased. In contrast, during application of 90 nA, the increase in the negative amplitude of NTIs occurred in an all-or-none manner. At 90 nA, NTIs that had the large increase in negative amplitude were averaged separately from those which did not show this effect.

Figure 3.

Effects of perfusing the recording electrode with zero Ca²⁺ solution (A) or low Na⁺ solution (B) on the hyperpolarization-induced changes in cold receptor NTI shape. (A and B) Averaged NTIs, from the same receptor, recorded before and during application of zero Ca²⁺ solution or low Na⁺ solution. In each case, NTIs were recorded in the absence of polarizing current and during application of +30 nA. The lower records in each panel show the control and treated (thick lines) NTIs overlaid. Between application of zero Ca²⁺ and low Na⁺ solution, the electrode was washed with control solution for 15 min. In this experiment, the low Na⁺ solution was prepared by replacing the NaCl with choline chloride.

Figure 4.

Effects of locally applied TTX (1 μM) on the polarization-induced changes in NTI shape in cold (A and B) and polymodal receptors (C and D). (A and C) Averaged NTIs record in single experiments before and during TTX application. In each case, NTIs were recorded in the absence of polarizing current and during application of hyperpolarizing (+ve) and depolarizing (−ve) current. The lower records in (A) and (C) show control and TTX (thick line) traces overlaid. (B and D) Graphs showing the mean effects of polarizing current on the positive (fill bars) and negative (open bars) amplitude of NTIs before and during application of TTX (cold receptors n = 9; polymodal receptors n = 9). In the graphs, the positive and negative amplitudes of NTIs were normalized to the positive amplitude measured in the absence of polarizing current (zero current). Statistical comparisons were made with paired sign tests. *, P < 0.05; **, P < 0.01.

Figure 5.

Effects of locally applied lidocaine (10 mM) on the polarization-induced changes in NTI shape in cold (A and B) and polymodal receptors (C and D). (A and C) Averaged NTIs recorded in single experiments before and during lidocaine application. In each case, NTIs were recorded in the absence of polarizing current and during application of hyperpolarizing (+ve) and depolarizing (−ve) current. The lower records in (A) and (C) show control and lidocaine (thick line) traces overlaid. (B and D) Graphs showing the mean effects of polarizing current on the positive (fill bars) and negative (open bars) amplitudes of NTIs before and during application of lidocaine (cold receptors n = 10; polymodal receptors n = 9). In the graphs, the positive and negative amplitudes of NTIs were normalized to the positive amplitude measured in the absence of polarizing current (zero current). Statistical comparisons were made with paired sign tests. *, P < 0.05; **, P < 0.01.

Figure 6.

Effects of locally applied TEA (20 mM) and 4-AP (1 mM) on polarization-induced changes in NTI shape in cold receptors. (A) Averaged NTIs recorded in the absence of polarizing current in a single experiments before and during application of TEA and 4-AP. The right hand records show the control and treated (thick line) traces overlaid. (B) Graphs showing the mean effects of polarizing current on the positive (fill bars) and negative (open bars) amplitude of NTIs before and during application of TEA and 4-AP (n = 9). In the graphs, the positive and negative amplitudes of NTIs were normalized with respect to the positive amplitude measured in the absence of polarizing current (zero current). Statistical comparisons were made with paired sign tests. *, P < 0.05; **, P < 0.01.