Calcitonin gene-related peptide suppresses hair cell responses to mechanical stimulation in the Xenopus lateral line organ

Abstract

The presence of calcitonin gene-related peptide (CGRP) in the efferent fibers of virtually every hair cell organ studied suggests it may serve some fundamental but heretofore unknown role in control of hair cell function. We examined the effects of CGRP on spontaneous and stimulus-evoked discharge patterns in an in vitro preparation of the lateral line organ of Xenopus laevis. Discharge patterns were determined by sinusoidally displacing the cupula with a glass micropipette driven with a piezoelectric device while recording afferent fiber activity. All afferent fibers had characteristic frequencies of 16-32 Hz. Responses synchronized to cupular displacements as small as 20 nm. CGRP suppressed responses of the lateral line organ to displacement while increasing spontaneous discharge rate. In the presence of CGRP, stimulus-response curves were shifted 10 dB toward higher displacement levels. The suppression of stimulus-evoked responses suggests a function for CGRP as an efferent neurotransmitter that is similar to that of cholinergic efferent transmission in other hair cell organs. The 10 dB shift toward larger displacements makes it comparable in magnitude with the effects of electrical stimulation of efferents in the mammalian cochlea. This suggests a significant role for CGRP in efferent modulation of the output of this mechanosensory organ.

Fig. 1.

Afferent nerve fibers of the lateral line organ discharge synchronously to mechanical movement of the cupula. Period histograms represent 10 presentations of sinusoidal cupular displacement at the characteristic frequency of the fiber. Voltage applied to the piezoelectric device is shown in the top. The bottom panels indicate responses in two different fibers to different displacement levels. At the lowest displacement levels (opencircles), responses were below the synchronization threshold of the fibers. At moderate displacement levels (filled gray circles), responses were synchronized to the stimulus waveform but below the rate threshold of the fibers. At the highest displacement levels (filled black circles), responses evoked increases in afferent discharge.

Fig. 2.

Both SI and DR increased with displacement. Data are summarized from 20 animals. Not all displacement levels were used in all experiments. Mean ± SE are plotted for displacements with three or more data points. DR was calculated by dividing the discharge rate during stimulus presentation by the discharge rate immediately after the stimulus presentation. Sigmoidal curves were fit to the SI data using a least squares regression analysis.

Fig. 3.

Threshold tuning curves for five fibers were generated from isodisplacement functions using SI (top) and DR (bottom) criterion levels for threshold. Threshold criteria for SI were displacements producing an SI of 0.15. Threshold criteria for DR were displacements producing a DR of 1.25. High (above 128 Hz) and low (below 1 Hz) frequency tails of the tuning curves are extrapolated from the isodisplacement functions by interpolation of the last two data points.

Fig. 4.

CGRP increased spontaneous discharge rate, an effect lasting long after CGRP was washed out. The preparation was continually perfused with a balanced salt solution. CGRP was administered at 10 μm during the time indicated with thebar.

Fig. 5.

Time course of the change in spontaneous rate, SI, and DR with CGRP perfusion. CGRP was applied at time 0 at concentrations of 5–50 μm. Each data point represents the average for 10 animals in which displacements were 2 μm or smaller and responses were stable for 50 min or more. Data points for each variable were taken once per minute.

Fig. 6.

CGRP suppresses response to mechanical stimulation. Period histograms before (filled circles) and after (open circles) CGRP are plotted for two representative fibers. The fiber in thetop was stimulated with a relatively low cupular displacement, whereas that in the bottom was stimulated with high displacement. Both fibers were stimulated at CF. For the fiber in the top, discharge rate was modulated around spontaneous without an overall increase in rate during stimulation. For the fiber in the bottom, displacements produced an increase in rate. Insets are examples (from a different fiber) of the change produced in the period histogram by lowering the gain of the stimulus by 6 dB.

Fig. 7.

CGRP shifts the displacement–response function to higher displacement levels. Data (mean ± SE) taken before (filled circles) and after (open circles) CGRP administration are plotted on the same axes. SI values before and after CGRP administrations were normalized to the sigmoidal curve from Figure 2. A sigmoidal curve was fit to all of the CGRP data (dashed line) by a least squares regression analysis. The curve was constrained to the normal maximum SI. There were three cases at 10 μm, six cases at 2 μm, three cases at 0.4 μm, two cases at 0.2 μm, and one case at 0.1 μm.

Fig. 8.

Our view of the effects of CGRP is conceptualized in this simulation of afferent discharge in response to a tone at which spontaneous rate is increased (30%) and the depth of modulation is reduced 50%.