Tonic inhibition alternates in paired neurons that set direction of fish escape reaction

Abstract

Crossed antagonism between activities in neurons subserving alternating movements such as swimming or walking has been described in a number of systems. The role of reciprocal inhibition has been implicated in these activities, but involvement of rhythmic ongoing fluctuations of membrane potential, called synaptic "noise," has not been examined. In the Mauthner (M) cells, which control the direction of escape, this activity is inhibitory. We report that in the zebrafish (Danio rerio), inhibitory synaptic noise exhibits prolonged bursts of rhythmic, inhibitory postsynaptic potentials, which attenuate the M cell's sensibility to excitatory sensory drives. Furthermore, paired intracellular recordings have shown that inhibitory synaptic noise alternates between two distinct states, noisy and quiet, which are out of phase in the two cells. Firing of either M cell resets this pattern by reducing the inhibition in the contralateral one. This suggests that an avoidance reflex in one direction may favor initiation, by the opposite M cell, of a subsequent escape toward a more appropriate location.

Figure 1

Alternate states of ISN. (A) Diagram of the experimental set-up and of M cells’ inhibitory networks. Action potentials were induced in the left (L) or the right (R) M cells by antidromic (AD) stimulation of the M axon(s) or by brief, intracellular current injections (≈1 msec) through the recording microelectrode. Both procedures activated the M cell ipsilateral recurrent pathway via the cranial relay neurons (crn), which synapse on collateral inhibitory interneurons (coll.). The commissural inhibitory interneurons (comm.), which terminate on both M cells, were activated by auditory stimuli via VIII nerve primary fibers. ac, axon cap. (▴ and ▵) Inhibitory and excitatory synapses, respectively. (B) Simultaneous intracellular recordings from both M cells. Bursts of fast IPSPs (arrows), present during the noisy state, disappear during the subsequent quiet period (crossed arrows). (C) Boxed region in B displayed at higher magnification and faster sweep speed.

Figure 2

Differential effects of M cell firing. (A and B) Superimposed (n = 3) intracellular recordings from a suppressive M cell. (A) A spinal stimulus (AD) produced a transient suppression of the ISN. (B) A spontaneous discharge in the zebrafish M cell increased the rate of spontaneously occurring IPSPs. (C and D) Recordings at a faster sweep speed, from the same cell as above, during a quiet period (n = 6 trials). (C) A spinal stimulation produced only an antidromic spike and a collateral IPSP. (D) A spontaneous discharge of the M cell was followed by an awakening of the ISN. (E) Simultaneous recordings at fast (Left) and slow (Right) sweep speeds from two M cells before and after a spinal stimulus (AD). The strength of the stimulation was adjusted to excite only the right cell (crossed arrow). A collateral IPSP (arrowheads) was induced in both cells but was followed by a transient burst of IPSPs in the activated neuron alone.

Figure 3

Reciprocal modulation of ISN. Simultaneous recordings from the left (L) and right (R) M cells and selective activation of one of them. (A and B) A directly evoked spike (truncated, arrowhead) followed by a collateral IPSP and a burst of noise were produced in the right (A) and left (B) M cells. ISN was suppressed simultaneously in the opposite side. (C) Simultaneous extracellular recordings from the right (Upper) and left (Lower) axon caps (circled areas) from another experiment. Note the spontaneous alternation of firing in both axon caps (crossed arrow) and after a spinal stimulation (AD), which presumably activated only the right M cell (arrow).

Figure 4

Properties of inhibitory responses and network model accounting for the alternation of ISN. (A) Spikes evoked by sound (500 Hz, 35-msec duration, 70 dB; arrow) extracellularly recorded in the left and right (L and R) axon caps (circled areas) were correlated (asterisks) in presumed commissural neurons (same experiment as in Fig. 3E). (B) Simultaneous failure to induce the collateral IPSP and ISN after direct activation (arrow) of the M cell at rates more than four to five per sec. (C) Typical rhythmic bursts (between brackets) recorded from another M cell. (D) Putative network (thick lines) accounting for the transition between states incorporated in the diagram of Fig. 1A (shaded here). Collateral interneurons (coll.) are inhibited by hypothetical, mutually connected crossed interneurons (ci), which can be activated by collaterals of the opposite M cell axon.

Figure 5

Temporal properties and inhibitory effects of ISN. (A Upper) Large periods of activity (between brackets) interrupted by faster waves. The autocorrelogram (Right) indicates a dominant rhythm of 60 Hz. (Lower) Bursts of IPSPs occurring at a frequency of 80 Hz in another cell. (B) Instantaneous frequencies of IPSPs in a simple ISN (Inset), plotted against time. M spikes (arrows) were evoked (n = 4 trials) by intracellular current injections that increased the frequency of the noise from 30 to 130 Hz. Control IPSPs were either continuous (n = 3, open symbols) or were induced during the quiet state (♦). (C1) Superimposed traces of antidromic M spikes (n = 10) recorded during the quiet (Left) or the noisy (Center) state and their superimposed averages (Right), indicating a 38% reduction during the noisy state (Right). (C2) Same presentation as above of excitatory postsynaptic potentials (arrow) followed by IPSPs (crossed arrow) evoked by auditory stimuli (sound, 500 Hz, 25 cycles, 50 dB). Note a 36% mean reduction during the noisy periods.