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Idiopathic hypersomnia (IH) is a chronic neurologic disorder with unknown mechanisms that result in long night-time sleep, daytime sleepiness, long non-refreshing naps, and difficult awakening presenting as sleep drunkenness. IH patients are typically diagnosed by shorter sleep latency on multiple sleep latency test (MSLT) along with long sleep time. Only symptomatic drug treatments are currently available for IH and no animal model to study it. Sleepy mice carry a splicing mutation in the Sik3 gene, leading to increased sleep time and sleep need. Here we used a mouse version of MSLT and a decay analysis of wake EEG delta power to validate the Sleepy mutant mouse as an animal model for IH. Sleepy mice had shorter sleep latency in the dark (active) phase than wild-type mice. They also showed lower decay of EEG delta density during wakefulness, possibly reflecting increased sleep inertia. These data indicate that the Sleepy mouse may have partial face validity as a mouse model for idiopathic hypersomnia. We then investigated the effect of orexin-A and the orexin receptor 2-selective agonist YNT-185 on the sleepiness symptoms of the Sleepy mouse. Intracerebroventricular orexin-A promoted wakefulness for 3 h and decreased wake EEG delta density after injection in Sleepy mice and wild-type mice. Moreover, Sleepy mice but not wild-type mice showed a sleep rebound after the orexin-A-induced wakefulness. Intraperitoneal YNT-185 promoted wakefulness for 3 h after injection in Sleepy mice, indicating the potential of using orexin agonists to treat not only orexin deficiency but hypersomnolence of various etiologies., Competing Interests: Declaration of competing interest None, (Copyright © 2023 Elsevier B.V. All rights reserved.)

We investigated the role of endothelin ETB receptor in the development of monocrotaline-induced pulmonary hypertension, by using the spotting-lethal (sl) rat, which carries a naturally occurring deletion in the endothelin ETB receptor gene. Three weeks after injection of saline or monocrotaline (60 mg/kg, s.c.), hemodynamics, cardiac hypertrophy and endothelin-1 levels in right ventricle were determined. Monocrotaline produced a marked pulmonary hypertension associated with increases in right ventricular pressure and hypertrophy, pulmonary arterial medial thickening and the endothelin-1 levels. The monocrotaline-induced alterations tended to be enhanced in ETB-deficient homozygous rats, compared with cases in wild-type rats. The treatment with selective ETA receptor antagonist ABT-627 [2R-(4-methoxyphenyl)-4S-(1,3-benzodioxol-5-yl)-1-(N,N-di(n-butyl)aminocarbonyl-methyl)-pyrrolidine-3R-carboxylic acid] for 3 weeks (10 mg/kg/day, twice daily) almost completely suppressed the monocrotaline-induced pulmonary hypertension and related organ damage both in ETB-deficient and wild-type animals to the same levels. Thus, we suggest that the antagonism of the ETA receptor is essential for the protection from monocrotaline-induced pulmonary hypertension, irrespective of the presence of the ETB receptors, although a protective role of ETB receptor-mediated action in the pathogenesis of this disease model cannot be ruled out.

We investigated the role of endothelin ET(B) receptor in the remnant kidney model of chronic renal failure, by using the spotting-lethal (sl) rat, which carries a naturally occurring deletion in the endothelin ET(B) receptor gene. After 5/6 nephrectomy, systolic blood pressure and renal functional parameters were measured for 12 weeks. At the end of the experimental period, arterial blood sample, remnant kidney, heart and aorta were collected and used for biochemical measurements and histopathological studies. The ET(B)-deficient sl/sl rats exhibited earlier and higher increases in systolic blood pressure, urinary protein excretion, blood urea nitrogen and plasma creatinine concentration, compared with cases in wild-type rats. Histopathologic examination of the kidney revealed glomerular and tubular lesions, alterations of which were more severe in sl/sl than in wild-type rats. While aortic endothelin-1 contents were increased similarly in both groups, the level of renal endothelin-1 content was significantly elevated in sl/sl rats, but not in the wild-type rats. These results suggest that enhanced endothelin-1 production is at least partly responsible for the increased susceptibility to partial ablation-induced chronic renal failure in ET(B) receptor-deficient rats and that ET(B) receptor-mediated actions are protective against vascular and renal injuries in this disease.

We determined the role of endothelin ET(B) receptor in the renal hemodynamic and excretory responses to big endothelin-1, using A-192621, a selective endothelin ET(B) receptor antagonist and the spotting-lethal (sl) rat, which carries a naturally occurring deletion in the endothelin ET(B) receptor gene. An intravenous injection of big endothelin-1 produced a hypertensive effect, which is greater in wild-type (+/+) rats pretreated with A-192621 and in homozygous (sl/sl) rats. Big endothelin-1 markedly increased urine flow, urinary excretion of sodium and fractional excretion of sodium in wild-type rats treated with the vehicle. These excretory responses to big endothelin-1 were markedly reduced by pharmacological endothelin ET(B) receptor blockade. On the other hand, big endothelin-1 injection to the endothelin ET(B) receptor-deficient homozygous animals resulted in a small diuretic effect. When renal perfusion pressure was protected from big endothelin-1-induced hypertension by an aortic clamp, the excretory responses in vehicle-treated wild-type rats were markedly attenuated. In homozygous or A-192621-treated wild-type rats, there was a small but significant decreasing effect in urine flow. In addition, big endothelin-1 significantly elevated nitric oxide (NO) metabolite production in the kidney of wild-type rats but not in the homozygous rats. We suggest that the diuretic and natriuretic responses to big endothelin-1 consist of pressure-dependent and pressure-independent effects and that the increased NO production via the activation of endothelin ET(B) receptors in the kidney is closely related to the big endothelin-1-induced excretory responses.

Pharmacological characteristics of [D-Pro9,[spiro-gamma-lactam]Leu10,Trp11]physalaemin-(1-11) (GR82334), a tachykinin NK1 receptor antagonist, and its effects on slow depolarizing responses of lumbar ventral roots evoked by primary afferent stimulation were examined in isolated spinal cord preparations of neonatal rats. GR82334 (1-3 microM) caused dose-dependent rightward shifts of the concentration-response curves for substance P, substance P methyl ester, delta-aminovaleryl [Pro9,N-Me-Leu10]substance P-(7-11) (GR73632) and neurokinin A in normal artificial cerebrospinal fluid and those for substance P methyl ester, GR73632 and neurokinin A in the presence of tetrodotoxin. GR82334 (10 microM) did not evoke gamma-aminobutyric acid (GABA) release from spinal cords of neonatal rats, whereas [D-Pro9,[spiro-gamma-lactam] Leu10,Trp11]substance P (GR71251), another tachykinin NK1 receptor antagonist, induced a significant increase in GABA release. GR82334 (1-3 microM) markedly depressed the slow depolarizing response of ventral roots, referred to as slow ventral root potential, evoked by the stimulation of the contralateral dorsal root or the ipsilateral saphenous nerve. In contrast, cyclo[Gln,Trp,Phe,Gly,Leu,Met] (L-659,877, 1 microM), a selective tachykinin NK2 receptor antagonist, did not depress the saphenous nerve-evoked slow ventral root potential and did not antagonize the action of neurokinin A to induce ventral root depolarization. The present results provide further evidence for the involvement of substance P, neurokinin A and tachykinin NK1 receptors in the primary afferent-evoked slow ventral root potentials.

We have developed an isolated spinal cord-skin preparation of the newborn rat. The spinal cord together with a piece of skin connected to the cord by the saphenous nerve was isolated from 1- to 4-day-old rats and separately superfused with artificial cerebrospinal fluid in two neighbouring chambers. Potentials were recorded extracellularly from the third lumbar ventral root. Application of capsaicin (0.5-2 microM) or KCl (60-350 mM) with brief pressure pulses to the perfusion bath of the skin evoked a depolarizing response of 20- to 40-s duration in the ventral root. The response was depressed by [Met5]enkephalin (0.03-3 microM), morphine (0.1-2 microM) and a tachykinin antagonist, [D-Arg1,D-Trp7,9,Leu11] substance P (spantide), (1-10 microM), applied to the spinal cord by superfusion, whereas the response was augmented by centrally administered calcitonin gene-related peptide (0.1-0.2 microM) or bicuculline (0.5-1 microM).

An in vitro preparation for testing antinociceptive drugs is described. The preparation consists of isolated spinal cord, spinal nerve roots and functionally connected tail of the newborn rat. Noxious pressure stimulation given to the tail induced in a lumbar ventral root a depolarizing response of 1-2 mV lasting about 15-30 s, which is referred to as tail-pinch potential. Single shock stimulation of a lumbar dorsal root induced in the ipsilateral ventral root of the same segment monosynaptic and polysynaptic reflexes, which were followed by a depolarizing response lasting ca. 20 s. The latter slow response is referred to as ipsilateral slow ventral root potential (VRP). Morphine, [Met5]enkephalin, [Leu5]enkephalin and [D-Ala2,Met5]enkephalinamide depressed both the tail-pinch potential and the ipsilateral slow VRP, but did not exert any noticeable effect on the monosynaptic and polysynaptic reflexes. The effects of morphine and enkephalins were completely abolished by naloxone. Naloxone potentiated the tail-pinch potential and the ipsilateral slow VRP in the fresh preparations which had not been exposed to opiate compounds or enkephalins, suggesting that the endogenous enkephalinergic mechanism occurs in the spinal cord and modulates the incoming nociceptive signals. The present isolated spinal cord-tail preparation will be useful for studying actions of analgesic drugs and for screening compounds for analgesic effects.

The role of the N-terminus in the activity of substance P analogues was studied with the isolated rat spinal cord and the guinea-pig ileum. When the C-terminal Leu10 or Met11 was replaced by the D-isomer, the analogues without the N-terminal tetrapeptide were 40-100 times less potent than those with the N-terminus in both motoneuron-depolarizing and ileum-contracting actions. These results that the N-terminus may stabilize the active conformation of the substance P analogues.

Endothelin, a 21-amino acid peptide produced by vascular endothelial cells, caused a sustained constriction of isolated large cerebral arteries of cats in a dose-dependent manner. The increased tone of the tissue did not return to the resting level after repeated washings. No vasodilator response was evoked by endothelin in the presence of an active tone. The contractile response of cerebral arteries was not inhibited by rubbing of the endothelium, cold storage denervation or indomethacin. In contrast, nicardipine or diltiazem antagonized the endothelin-induced contraction non-competitively. No contraction was evoked by endothelin in a Ca2+-free solution while the addition of Ca2+ ions in the presence of endothelin in a Ca2+-free solution caused a sustained contraction. Ca2+-induced contraction in the Ca2+-free solution containing endothelin was also inhibited by nicardipine. Therefore, endothelin causes a direct contraction of the smooth muscles of cat cerebral arteries, probably by activating the influx of Ca2+ ions through L-type Ca2+ channels of smooth muscles.