Sedation enhances the following frequency of the GF pathway and is dependent on a functional slo gene. Frequency-response curves (Left) and the FF50 plots (Right) for (A) wild-type flies that were sedated with benzyl alcohol (BA) 24 h earlier and for the nonsedated controls (Control), (B) ash2¹⁸/slo⁴ transheterozygotes that do not express slo in the nervous system and that were sedated with benzyl alcohol (BA) 24 h earlier and for the nonsedated controls (Control), (C) B52H flies carrying a heat-shock inducible slo cDNA transgene that were heat-shocked (slo induced) for 30 min 24 h earlier and for the nonheat-shocked controls (Control), and (D) for the parental non transgenic stock (w¹¹¹⁸) that were heat-shocked (Mock-induced) for 30 min 24 h earlier and for the nonheat-shocked controls (Control). Error bars are SEM, n > 7 each; **P < 0.05 by repeated measures ANOVA; *P < 0.05 by Student's t test.

Behavioral and electrophysiological manifestation of tolerance. Flies were sedated with benzyl alcohol in a vapor chamber for 20 min and immediately tested for behavioral recovery, following frequency, and seizure susceptibility. (A) Behavioral recovery plot for flies recovering from the first and second exposure (first exposure was 24 h prior) to benzyl alcohol in the vapor chamber. (B) Average time for return of righting reflex after benzyl alcohol exposure. (C) Capacity of the GF to follow a 200-Hz stimulation, 2 min after the 20-min sedation period. Number of successful responses from 30 stimuli were counted in awake, wild-type flies, in flies recovering from their first exposure, and in flies recovering from their second exposure. (D) Benzyl alcohol concentration per fly immediately after the first and second exposures. (E) Seizure activity plots of flies after the first and second exposure to benzyl alcohol in the vapor chamber. Seizures were evoked by a 50 V ECS every 10 min, starting after a 2-min mounting delay after removal from the vapor chamber. (F) Average time for return of seizure response. n = 9 each; **P < 0.01 by Student's t test, *P < 0.05 by one-way ANOVA, Newman-Keuls multiple comparison test. All error bars are SEM.

The GF pathway. (A) GF neurons receive input from visual or other neural afferent pathways (Aff.) and are electrically connected to the peripheral synapsing interneuron (PSI) that synapses on the DLM motoneurons (DLMmn). The DLMmn synapses on the DLM. Stimulating potentials through electrodes on the eyes activate neural afferent pathways and trigger a response that can be recorded by an electrode in the DLM. (B) Representative traces of short-latency recordings from the DLM after 100-Hz and 160-Hz stimuli. Successful DLM responses closely follow each stimulus artifact at 100 Hz but fail at higher frequency stimulation. (Scale bars: 10 ms, 5 mV.) (C) Number of successful responses are plotted against frequency after GF stimulation (GF Pathway) or direct-muscle stimulation (Muscle) via electrodes in the thoracic ganglion. Note that the capacity of the muscle to follow high-frequency stimulations far exceeds that of the GF pathway. Error bars are SEM, n = 5 each.

Sedation and increased slo expression increase seizure susceptibility. Electroconvulsive stimuli (S) of varying voltages were applied to determine seizure threshold of flies after different treatments. A constant low-frequency stimulus was applied throughout the duration of the recording to assess the responsiveness of the GF pathway. (A) A typical seizure repertoire consisting of a high-frequency initial discharge (ID) followed by a period of evoked response failures (Failure), a delayed discharge (DD), and recovery from response failures. Seizure threshold curves (Left) and average seizure threshold (Right) are shown for (B) wild-type flies that were sedated with benzyl alcohol (BA) 24 h earlier and for the nonsedated controls (Control), (C) B52H flies carrying a heat-shock inducible slo cDNA transgene that were heat-shocked (slo induced) 24 h earlier and for the nonheat-shocked controls (Control), and (D) for the parental non transgenic stock (w¹¹¹⁸) that were heat-shocked (Mock-induced) 24 h earlier and for the nonheat-shocked controls (Control). Error bars are SEM, n > 9 each; *P < 0.05 by Student's t test.

Schematic model of a slo-mediated homeostatic response underlying drug tolerance and a withdrawal phenotype. (A) In a “no drug” state, a balance between excitation and inhibition of neural activity in the brain prevails, allowing for normal physiological and behavioral activity, including average seizure susceptibility to electroconvulsive shock (Δ). (B) During exposure to a sedative drug, this balance is pushed toward a more inhibited state characterized by sedation, decreased following frequency, and suppression of seizures. Drug exposure also produces an increase in slo expression. (C) After drug withdrawal, the increase in slo expression is unmasked, resulting in an excited neural state and characterized by increased neuronal following frequency and higher seizure susceptibility. (D) When drug exposure is repeated, the slo-mediated compensation resists the effect of the drug, resulting in a milder reduction in following frequency and reduced seizure suppression. The electrophysiological traces shown here are schematic representations of hypothetical data, not real traces.