No abstract

The cellular mechanisms underlying the physiological eects of the orexins are poorly understood. Therefore, the pharmacology of the recombinant human orexin receptors was studied using FLIPR. Intracellular calcium ([Ca 2+ ] i ) was monitored in Chinese hamster ovary (CHO) cells stably expressing orexin-1 (OX 1 ) or orexin-2 (OX 2 ) receptors using Fluo-3AM. Orexin-A and orexin-B increased [Ca 2+ ] i in a concentration dependent manner in CHO-OX 1 (pEC 50 =8.03+0.08 and 7.30+0.08 respectively, n=5) and CHO-OX 2 (pEC 50 =8.18+0.10 and 8.43+0.09 respectively, n=5) cells. This response was typi®ed as a rapid peak in [Ca 2+ ] i (maximal at 6 ± 8 s), followed by a gradually declining secondary phase. Thapsigargin (3 mM) or U73122 (3 mM) abolished the response. In calcium-free conditions the peak response was unaected but the secondary phase was shortened, returning to basal values within 90 s. Calcium (1.5 mM) replacement restored the secondary phase. In conclusion, orexins cause a phospholipase C-mediated release of calcium from intracellular stores, with subsequent calcium in¯ux.

The link between the cognitive deficit associated with Alzheimer type dementia and the loss of cholinergic function in the disease provides a basis for examining muscarinic agonists as potential therapeutic agents. This paper describes the design and synthesis of novel azabicyclic methyl esters as ligands for the muscarinic receptor. Replacement of the methyl ester by a 3-methyl-1,2,4-oxadiazole ring produces potent metabolically more stable muscarinic agonists capable of penetrating the central nervous system. These compounds generally show improved affinity relative to the corresponding methyl esters. 3-Methyl-1,2,4-oxadiazole 7b has an affinity 4 times that of acetylcholine. Receptor affinity is discussed in relation to the size and geometry of the azabicyclic ring and the electronic properties of the heteroaromatic ring.

The synthesis of 15 methyl or unsubstituted 1,2,3-triazoles, 1,2,4-triazoles, and tetrazoles additionally substituted with a 1-azabicyclo[2.2.2]octan-3-yl group is described. The potency and efficacy of these compounds as muscarinic ligands were determined in radioligand binding assays using [3H]oxotremorine and [3H]quinuclidinyl benzilate. Potency and efficacy were found in compounds in which the azole moiety was attached to the azabicyclic ring either through a carbon atom or a nitrogen atom. Electrostatic potential maps of both the C-linked and the novel N-linked series of compounds were calculated. A relationship between position and depth of the electrostatic minima relative to the azabicyclic ring and the potency and efficacy of the compounds was determined.

1 This study describes the pharmacological comparison of the muscarinic partial agonists sabcomeline, xanomeline and milameline at human cloned muscarinic receptor subtypes (hM 1 ± 5 ). 2 Radioligand binding studies at the hM 1 ± 5 muscarinic receptor subtypes were compared with functional studies using microphysiometry using carbachol as the standard full agonist. 3 In binding assays none of the compounds studied displayed preferential a nity for the M 1,3,4 or M 5 subtypes although carbachol was less potent at hM 1 than hM 3,4,5 . 4 In functional studies, all of the compounds studied displayed similar levels of e cacy across the muscarinic receptors with the exception of M 3 , where there was a large apparent receptor reserve and the compounds behaved essentially as full agonists. 5 Sabcomeline was the most potent agonist in functional studies but also showed the lowest e cacy. In terms of potency, xanomeline showed some selectivity for M 1 over M 2 receptors and milameline showed some selectivity for M 2 over M 1 receptors. 6 These results show the value of microphysiometry in being able to compare receptor pharmacology across subtypes irrespective of the signal transduction pathway. 7 None of the partial agonists showed functional selectivity for M 1 receptors, or indeed any muscarinic receptor, in the present study.