AN ADENOSINE A2a AGONIST INCREASES SLEEP AND INDUCES FOS IN VENTROLATERAL PREOPTIC NEURONS T. E. SCAMMELL, a; 1 * D. Y. GERASHCHENKO, b; 1 T. MOCHIZUKI, b M. T. MCCARTHY, a I. V. ESTABROOKE, a C. A. SEARS, a C. B. SAPER, a;d Y. URADE b;c and O. HAYAISHI b a Department of Neurology, Beth Israel Deaconess Medical Center, 77 Avenue Louis Pasteur, Boston, MA 02115, USA b Osaka Bioscience Institute, Osaka, Japan c Core Research for Evolutional Science and Technology, Japan Science and Technology Corporation, Osaka, Japan d Program in Neuroscience, Harvard Medical School, Boston, MA, USA AbstractöConsiderable evidence indicates that adenosine may be an endogenous somnogen, yet the mechanism through which it promotes sleep is unknown. Adenosine may act via A1 receptors to promote sleep, but an A2a receptor antagonist can block the sleep induced by prostaglandin D 2 . We previously reported that prostaglandin D 2 activates sleep-promoting neurons of the ventrolateral preoptic area, and we hypothesized that an A2a receptor agonist also should activate these neurons. Rats were instrumented for sleep recordings, and an injection cannula was placed in the sub- arachnoid space just anterior to the ventrolateral preoptic area. After an 8^10-day recovery period, the A2a receptor agonist CGS21680 (20 pmol/min) or saline was infused through the injection cannula, and the animals were killed 2 h later. The brains were stained using Fos immunohistochemistry, and the pattern of Fos expression was studied in the entire brain. CGS21680 increased non-rapid eye movement sleep and markedly increased the expression of Fos in the ventrolateral preoptic area and basal leptomeninges, but it reduced Fos expression in wake-active brain regions such as the tuberomammillary nucleus. CGS21680 also induced Fos in the shell and core of the nucleus accumbens and in the lateral subdivision of the central nucleus of the amygdala. To determine whether these e¡ects may have been mediated through A1 receptors, an additional group of rats received subarachnoid infusion of the A1 receptor agonist N 6 -cyclo- pentyladenosine (2 pmol/min). In contrast to CGS21680, infusion of N 6 -cyclopentyladenosine into the subarachnoid space produced only a small decrease in rapid eye movement sleep, and the pattern of Fos expression induced by N 6 -cyclopentyladenosine was notable only for decreased Fos in regions near the infusion site. These ¢ndings suggest that an adenosine A2a receptor agonist may activate cells of the leptomeninges or nucleus accumbens that increase the activity of ventrolateral preoptic area neurons. These ventrolateral preoptic area neurons may then coordinate the inhibition of multiple wake-promoting regions, resulting in sleep. ß 2001 IBRO. Published by Elsevier Science Ltd. All rights reserved. Key words: sleep, prostaglandin D 2 , ventrolateral preoptic area, tuberomammillary nucleus, somnogen, leptomeninges. Adenosine has been hypothesized to play a critical role in the production of sleep. Extracellular concentrations of adenosine in the basal forebrain increase during wake- fulness and decrease during sleep (Porkka-Heiskanen et al., 1997). Adenosine and adenosine agonists can in£u- ence the activity of putative state-regulatory neurons (Alam et al., 1999; Rainnie et al., 1994), yet it remains unclear whether adenosine's sleep-promoting e¡ects are mediated via adenosine A1 or A2a receptors. Adenosine may promote sleep by inhibiting the activity of wake- generating systems or by facilitating the activity of sleep-promoting systems. One essential population of sleep-promoting neurons is found in the ventrolateral preoptic area (VLPO). These cells have increased ¢ring rates during rapid eye movement (REM) and non-REM (NREM) sleep (Szymusiak et al., 1998) and are more active during sleep as indicated by the expression of Fos (Scammell et al., 2000; Sherin et al., 1996). These VLPO neurons appear necessary for the production of sleep because excitotoxic lesions of the VLPO region markedly decrease both NREM and REM sleep (Lu et al., 2000). The VLPO sends inhibitory GABAergic and galaninergic projections to many wake-promoting regions such as the histaminergic neurons of the tuber- omammillary nucleus (TMN) (Sherin et al., 1998). Through these projections, the VLPO may play a critical role in coordinating the inhibition of multiple arousal systems, thus promoting sleep. 653 1 These authors contributed equally to this work. *Corresponding author. Tel. : +1-617-667-0833 ; fax : +1-617-667- 0810. E-mail address : tscammel@caregroup.harvard.edu (T. E. Scammell). Abbreviations : Acb, nucleus accumbens; BSTLD, laterodorsal sub- nucleus of the bed nucleus of the stria terminalis; CeL, lateral subdivision of the central nucleus of the amygdala ; CPA, N 6 - cyclopentyladenosine ; IR, immunoreactive ; NREM, non-REM ; PBcl, central lateral subnucleus of the parabrachial nucleus ; PBel, external lateral subnucleus of the parabrachial nucleus; PBS, phosphate-bu¡ered saline ; PGD 2 , prostaglandin D 2 ; PVH, paraventricular nucleus of the hypothalamus ; REM, rapid eye movement ; SON, supraoptic nucleus ; TMN, tubero- mammillary nucleus ; VLPO, ventrolateral preoptic area. NSC 5224 7-11-01 www.neuroscience-ibro.com Neuroscience Vol. 107, No. 4, pp. 653^663, 2001 ß 2001 IBRO. Published by Elsevier Science Ltd Printed in Great Britain. All rights reserved PII:S0306-4522(01)00383-9 0306-4522 / 01 $20.00+0.00