Hypothalamic arousal regions are activated during modafinil-induced wakefulness

Abstract

Modafinil is an increasingly popular wake-promoting drug used for the treatment of narcolepsy, but its precise mechanism of action is unknown. To determine potential pathways via which modafinil acts, we administered a range of doses of modafinil to rats, recorded sleep/wake activity, and studied the pattern of neuronal activation using Fos immunohistochemistry. To contrast modafinil-induced wakefulness with spontaneous wakefulness, we administered modafinil at midnight, during the normal waking period of rats. To determine the influence of circadian phase or ambient light, we also injected modafinil at noon on a normal light/dark cycle or in constant darkness. We found that 75 mg/kg modafinil increased Fos immunoreactivity in the tuberomammillary nucleus (TMN) and in orexin (hypocretin) neurons of the perifornical area, two cell groups implicated in the regulation of wakefulness. This low dose of modafinil also increased the number of Fos-immunoreactive (Fos-IR) neurons in the lateral subdivision of the central nucleus of the amygdala. Higher doses increased the number of Fos-IR neurons in the striatum and cingulate cortex. In contrast to previous studies, modafinil did not produce statistically significant increases in Fos expression in either the suprachiasmatic nucleus or the anterior hypothalamic area. These observations suggest that modafinil may promote waking via activation of TMN and orexin neurons, two regions implicated in the promotion of normal wakefulness. Selective pharmacological activation of these hypothalamic regions may represent a novel approach to inducing wakefulness.

Fig. 1.

Effects of modafinil on rats treated at midnight. Modafinil produces clear increases in waking, whereas rats treated with vehicle have no increase in waking. *p < 0.001.

Fig. 2.

A series of line drawings illustrating the distribution of Fos-IR neurons in individual rats after administration of vehicle (A) or modafinil (75 mg/kg;B) at midnight. Modafinil-treated rats have substantial increases in Fos-IR neurons in hypothalamic arousal regions such as theTMN and PFx, as well as in theBSTLD and the CeL. Increased numbers of Fos-IR neurons can also be seen in the cortex, islands of Calleja (IC), and striatum, but not in the SCN orAHA.

Fig. 3.

Effects of vehicle (A, C, E) or modafinil (B, D, F) on Fos expression in putative sleep/wake regulatory regions. Administration of modafinil (150 mg/kg) at midnight substantially increases Fos expression in the TMN, but little change is evident in the SCN or AHA. Scale bars, 100 μm.3V, Third ventricle; ox, optic chiasm.

Fig. 4.

Few orexin-IR neurons contain Fos in rats treated with vehicle at midnight (A), but Fos-IR nuclei are common in orexin-IR neurons of rats treated with modafinil (150 mg/kg; B). Single arrow, Orexin-IR neuron; double arrows, Fos-IR and orexin-IR neuron. Scale bar, 30 μm.

Fig. 5.

Photomicrographs of several regions with notable increases in Fos-IR neurons after treatment with modafinil (150 mg/kg) at midnight. A, The IC have marked increases in the number of Fos-IR cells, but Fos expression is considerably less in other parts of the olfactory tubercle.B, The striatum has large increases in Fos expression.C, D, The BSTLD (C) and CeL (D) also show intense Fos expression that is not seen in other parts of the extended amygdala.E, F, Modafinil also induces Fos immunoreactivity at the mesopontine junction in the retro-ventral tegmental area (retro-VTA; E) and in theLC (F). Scale bars, 100 μm.ac, Anterior commissure; cc, corpus callosum; IPN, interpeduncular nucleus;LV, lateral ventricle; opt, optic tract;4V, fourth ventricle.

Fig. 6.

Effects of modafinil on rats treated at noon on a 12:12 hr LD cycle. Modafinil markedly increases waking, whereas vehicle has no effect. *p < 0.01.

Fig. 7.

Effects of modafinil on rats housed in constant dark and treated at noon. Rats treated with vehicle have a small and transient increase in waking, but those treated with modafinil have a dose-dependent increase in waking. *p < 0.01.