2. Effect of the application of GABA and glycine antagonists on the activity of the rat locus coeruleus neurons during sleep

Like in the study of Aston-Jones and Bloom (1981), noradrenergic LC neurons were identified by their typical broad action potentials (1.5-2ms duration), and a phasic excitatory response to a sensory stimulus (auditive click) immediately followed by an inhibition (Chouvet et al., 1988, Akaoka et al, 1992; Maloney et al., 1997). Their mean discharge rate was of 1.39 Hz during quiet W. During SWS, LC cells showed a decrease in their firing rate (0.56 Hz) while during PS episodes they were nearly quiescent (0.01 Hz) showing only occasional single spikes (Gervasoni et al., 1998).

Iontophoretic ejections of strychnine induced a sustained increase in the discharge rate of LC neurons regardless of the vigilance state: from 1.70 Hz to 11.41 during W, 0.39 to 9.34 during SWS (Fig. 2A) and 0.06 Hz to 14.13 Hz during PS (Fig. 2B)(Darracq et al., 1996). For some neurons, the rat displayed successive short periods of SWS and W during the effect of strychnine. In these neurons, we compared their increase in discharge rate during two successive SWS and W periods. We found that their increase of activity was 1,93 time superior during W that during SWS periods. This difference was statistically significant.

Like in anesthetized animals, (Luppi et al., 1991, Ennis and Aston-Jones, 1989) iontophoretic applications of glycine or GABA suppressed the spontaneous discharge of LC neurons during W or SWS, and glycine- but not GABA-induced inhibitions were antagonized by co-iontophoresis of strychnine.

Iontophoretic ejections of bicuculline during W, SWS and PS induced a progressive and sustained increase of the discharge rate of LC neurons without inducing a change in the vigilance state (Gervasoni et al., 1998). The firing frequency increased from 1.09 Hz to 8.72 Hz during W (Fig. 3B) and from 0.34 Hz to 6.91 Hz during SWS (Fig.3A). During PS, while LC neurons were practically silents (0.01 Hz), they showed following bicuculline application a remarkable firing rate of 6.59 Hz (Fig.3A,B). For some neurons, the rat displayed successive short periods of SWS and W during the effect of bicuculline. In these neurons, we compared their increase in discharge rate during two successive SWS and W periods. We found that their increase of activity was 1,23 time superior during W that during SWS periods. In contrast with strychnine, this difference was not statistically significant.

During W or SWS, GABA-induced inhibitions were fully antagonized by bicuculline co-iontophoresis, while GLY-induced inhibitions were unaffected (Fig.3).

Figure 2

Figure 2 : A : Effect of an iontophoretic application of strychnine (90nA). This application induced an activation of the neuron during SWS (EEG with high voltage slow activity and spindles), from 0.16 to 8,9 Hz starting 42 seconds after the onset of the drug application. During the following short period of W 193-200 s), the discharge rate of the neuron reached 14 Hz. Then, with the return of slow waves, it strongly decreases up to the next W phase (235-250s) during which the discharge rate of the neuron re-increase largely above the normal W values. The persistence of a decrease of activity during SWS of this LC cells during the strychnine effect suggest that another inhibitory neurotransmitter that GLY specifically inhibits LC cells during SWS.

Figure 2 : B : Effect of an iontophoretic application of strychnine during PS (150nA, 50s). Note the sustained activation of the LC unit starting about 40s after the onset of the strychnine application.

Figure 3

Figure 3 : A : Effects of two successive iontophoretic applications of bicuculline (100nA, 20s) during SWS and PS (characterized by flat EMG and desynchronized EEG). Note the small increase in the discharge rate of the LC neuron, from 0.6 to 1.8 Hz during SWS, and 0 to 3.6 Hz during PS. The effect appeared around 25 seconds after the onset of the ejections. The increase in firing rate did not alter the vigilance state of the animal.

Figure 3 : B : Polygraphic recordings displaying the EMG, EEG, the unit activity of a LC neuron, its firing rate, and the effect of the iontophoresis of bicuculline (50nA, 52s) during PS. GLY still inhibits this LC neuron during the bicuculline-induced activation. Note that during the effect of bicuculline, the firing rate of the LC cell did not further increase when the rat woke up (arrow). This result suggest that GABA is responsible for the inhibition of LC cells during PS.

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