Role of intrinsic conductances underlying responses to transients in octopus cells of the cochlear nucleus

Abstract

Recognition of acoustic patterns in natural sounds depends on the transmission of temporal information. Octopus cells of the mammalian ventral cochlear nucleus form a pathway that encodes the timing of firing of groups of auditory nerve fibers with exceptional precision. Whole-cell patch recordings from octopus cells were used to examine how the brevity and precision of firing are shaped by intrinsic conductances. Octopus cells responded to steps of current with small, rapid voltage changes. Input resistances and membrane time constants averaged 2.4 MOmega and 210 microseconds, respectively (n = 15). As a result of the low input resistances of octopus cells, action potential initiation required currents of at least 2 nA for their generation and never occurred repetitively. Backpropagated action potentials recorded at the soma were small (10-30 mV), brief (0.24-0.54 msec), and tetrodotoxin-sensitive. The low input resistance arose in part from an inwardly rectifying mixed cationic conductance blocked by cesium and potassium conductances blocked by 4-aminopyridine (4-AP). Conductances blocked by 4-AP also contributed to the repolarization of the action potentials and suppressed the generation of calcium spikes. In the face of the high membrane conductance of octopus cells, sodium and calcium conductances amplified depolarizations produced by intracellular current injection over a time course similar to that of EPSPs. We suggest that this transient amplification works in concert with the shunting influence of potassium and mixed cationic conductances to enhance the encoding of the onset of synchronous auditory nerve fiber activity.

Fig. 1.

Morphology of an octopus cell labeled by intracellular perfusion with biocytin and reconstructed with a camera lucida from a series of sections cut in the same coronal plane as the original slice. A labeled octopus cell is located in the most caudal and dorsal part of the posteroventral cochlear nucleus (PVCN). The recording was made from the cell body near the caudal surface of the slice. The dendrites of the cell extend anteriorly through the thickness of the slice. Its axon exits through a fiber tract that lies at the medial surface of the dorsal cochlear nucleus and goes around the inferior cerebellar peduncle (ICP) and spinal trigeminal tract (STT). A collateral innervates granule cell regions (Gr) that lie adjacent to the octopus cell area. The molecular layer (ML), fusiform cell layer (FCL), and deep layer (DL) of the dorsal cochlear nucleus overlie the PVCN.

Fig. 2.

Electrophysiological properties of octopus cells.A, Left, Responses of an octopus cell to steps of current between 2.8 and −2.8 nA in 0.4 nA increments. Responses to depolarizing current pulses are characterized by strong outward rectification. Inward rectification is visible in responses to hyperpolarizing current pulses as a delayed depolarizing sag. A, Right, Action potentials were small (14 mV) and narrow (0.3 msec base width) and could be evoked at the onset and offset of the large depolarizing and hyperpolarizing current pulses, respectively. Subthreshold responses to depolarizing current show a prominent “hump” at their onset (e.g., arrow).B, The voltage–current relationship of the same cell measured at the peak and steady-state of the responses shown inA (circles and squares, respectively). The steady-state input resistance within −5 mV of rest was 2.4 MΩ. C, The membrane time constant of the cell shown in A, measured from a 1.4 mV hyperpolarizing pulse, was described by a single exponential function (dark trace) and was 0.16 msec.

Fig. 3.

Action potentials in octopus cells.A, A large depolarizing current step evoked a small spike in an octopus cell that was blocked reversibly by 2 μm TTX. Current steps in Control and in2 μm TTX were 2.0 nA; inWash it was 2.4 nA. B, The amplitude of action potentials increased with the size of depolarizing current steps. C, Plot of spike amplitude versus the rate of change of the initial 120 μsec of the response shows that spike amplitude increased approximately linearly with the rate of change of voltage before saturation.

Fig. 4.

Amplification of subthreshold depolarizing responses by voltage-gated sodium and calcium channels.A, The initial response of an octopus cell to 0.9 and 1.9 nA depolarizing current pulses shows a depolarizing transient (a “hump”). In the presence of TTX the initial transients were reduced by ∼2 mV, indicating that voltage-sensitive Na⁺conductances had contributed to the transient response. Both larger and smaller responses were reduced further in a TTX/0 calcium solution, indicating an additional contribution from voltage-gated calcium channels. B, Amplitude of transient peak depolarizations versus stimulus current for the cell for which the traces are shown inA. Amplification of the initial voltage transient by sodium channels is apparent in responses >8 mV from rest, whereas amplification by calcium channels is apparent in responses >5 mV from rest. On elimination of the amplifying effects of sodium and calcium channels, an additional 1.1 nA of current was required to depolarize the cell to a level that was just below the action potential threshold in control conditions.

Fig. 5.

The effect of external cesium on the responses of an octopus cell to steps of injected current. A, External cesium (15 mm) decreased the current required for action potential initiation, reduced the delayed depolarizing sag in responses to hyperpolarizing current steps, and increased the input resistance of the cell, particularly in the hyperpolarizing voltage range. Resting potentials (in mV): Control, −61;15 mm Cs⁺, −66;Wash, −63. Current pulses: from 1.4 to −1.4 nA in −0.2 nA steps. B, The voltage–current relation derived from the traces in A. C, The membrane time constant of the cell in normal saline and 15 mmcesium. Hyperpolarizing responses of comparable magnitude in the two conditions were fit by single exponential functions (darker traces). The time constant increased by over an order of magnitude in the presence of external cesium. D, External cesium increased both the size and width of the action potential modestly and reversibly decreased the current necessary for their initiation from 2 to 1.6 nA.

Fig. 6.

Sodium dependence of inward rectification.A, Responses of an octopus cell to hyperpolarizing current steps of different magnitudes in normal physiological saline and saline in which extracellular sodium was reduced from 130 to 20 mm. The reduction of extracellular sodium hyperpolarized the cell and reduced the sag toward rest. B, Steady-state voltage–current relationship of the same cell shows an increase in the input resistance over the hyperpolarizing voltage range. C, The difference between the steady-state and peak voltages (indicated in A by arrows) in three different cells during the response to a −4.0 nA current step in normal and reduced extracellular sodium shows a statistically significant reduction in the mean sag in the presence of low extracellular sodium (Student’s t test;p < 0.05; n = 3).

Fig. 7.

The influence of 4-AP on the responses of an octopus cell to steps of injected current. A, Responses of an octopus cell to current pulses between 3.5 and −3.5 nA in −0.5 nA steps. In the presence of 4-AP the input resistance of the cell increased over both the positive and negative voltage ranges, and complex spikes were elicited. B, The voltage–current relationship for the traces shown in A. Voltage measurements are the difference between the average membrane potential during the last 1 msec of the response and the resting potential (−60 mV in control; −59 mV in 4-AP). C, The membrane time constant of the cell in the presence of 5 mm 4-AP increased by over an order of magnitude as compared with control (normal saline).D, The effect of 4-AP on the initiation and shape of action potentials reveals two components. In the presence of 4-AP the action potential was broadened and was evoked with smaller injected currents. Larger injected currents in the presence of 4-AP evoked a second inflection and a second, larger spike.

Fig. 8.

Calcium spikes evoked in the presence of internal cesium. In normal physiological saline the cell responded to a 3.4 nA current pulse with a small depolarizing hump, followed by a train of larger, wider spikes. Removal of extracellular calcium reversibly eliminated the large spikes, revealing their calcium dependence. The removal of calcium from the bathing medium also reduced the amplitude of the small transient at the onset of the response, indicating that it was mediated in part by a calcium current.