M1 muscarinic receptor is a key target of neuroprotection, neuroregeneration and memory recovery by i-Extract from Withania somnifera.

Memory loss is one of the most tragic symptoms of Alzheimer's disease. Our laboratory has recently demonstrated that 'i-Extract' of Ashwagandha (Withania somnifera) restores memory loss in scopolamine (SC)-induced mice. The prime target of i-Extract is obscure. We hypothesize that i-Extract may primarily target muscarinic subtype acetylcholine receptors that regulate memory processes. The present study elucidates key target(s) of i-Extract via cellular, biochemical, and molecular techniques in a relevant amnesia mouse model and primary hippocampal neuronal cultures. Wild type Swiss albino mice were fed i-Extract, and hippocampal cells from naïve mice were treated with i-Extract, followed by muscarinic antagonist (dicyclomine) and agonist (pilocarpine) treatments. We measured dendritic formation and growth by immunocytochemistry, kallikrein 8 (KLK8) mRNA by reverse transcription polymerase chain reaction (RT-PCR), and levels of KLK8 and microtubule-associated protein 2, c isoform (MAP2c) proteins by western blotting. We performed muscarinic receptor radioligand binding. i-Extract stimulated an increase in dendrite growth markers, KLK8 and MAP2. Scopolamine-mediated reduction was significantly reversed by i-Extract in mouse cerebral cortex and hippocampus. Our study identified muscarinic receptor as a key target of i-Extract, providing mechanistic evidence for its clinical application in neurodegenerative cognitive disorders.

A pharmacological evidence for the presence of antihistaminic and anticholinergic activities in Equisetum debile Roxb.

OBJECTIVE: The study was designed to evaluate possible antihistaminic and anticholinergic activities of Equisetum debile. MATERIALS AND METHODS: Effects of crude ethanolic (Ed.Eth) and effects of crude aqueous (Ed.Aq) extracts of E. debile were studied using isolated guinea pig ileum, rabbit jejunum, and rabbit trachea. Tissue responses were recorded using isotonic and isometric transducers, connected with PowerLab data acquisition system. RESULTS: A dose-dependent (0.1-0.3 mg/ml) rightward shift was demonstrated in histamine concentration-response curves. Whereas a complete relaxation of carbachol (1 µM)-induced contractions in isolated rabbit jejunum (3 mg/ml) and tracheal (10 mg/ml) preparations was observed, similar to dicyclomine at 1 and 3 µM, respectively. However, no significant difference between the effects of Ed.Eth and Ed.Aq was observed. CONCLUSION: Study provides pharmacological evidence for the presence of antihistaminic and anticholinergic activities in crude extracts of E. debile and also highlight its medicinal significance in the management of airway and gastrointestinal disorders.

Studies on antidiarrheal and antispasmodic activities of Lepidium sativum crude extract in rats.

This study was aimed to provide the pharmacological basis for the medicinal use of Lepidium sativum in diarrhea using in vivo and in vitro assays. The seed extract of Lepidium sativum (Ls.Cr) at 100 and 300 mg/kg inhibited castor oil-induced diarrhea in rats. In isolated rat ileum, Ls.Cr (0.01-5 mg/mL) reversed carbachol (CCh, 1 µM) and K(+) (80 mM)-induced contractions with higher potency against CCh, similar to dicyclomine. Preincubation of rat ileum with a lower concentration of Ls.Cr (0.03 mg/mL) caused a rightward parallel shift in the concentration-response curves (CRCs) of CCh without suppression of the maximum response, while at the next higher concentration (0.1 mg/mL), it produced a non-parallel rightward shift with suppression of the maximum response, similar to that of dicyclomine. Ls.Cr shifted the CRCs of Ca(++) to the right with suppression of the maximum response, similar to verapamil. These data suggest that Lepidium sativum seed extract possesses antidiarrheal and spasmolytic activities mediated possibly through dual blockade of muscarinic receptors and Ca(++) channels, though additional mechanism(s) cannot be ruled out and this study explains its medicinal use in diarrhea and abdominal cramps.

A computer-automated touchscreen paired-associates learning (PAL) task for mice: impairments following administration of scopolamine or dicyclomine and improvements following donepezil.

RATIONALE: Performance on the Cambridge Neuropsychological Test Automated Battery touchscreen paired-associates learning (PAL) test is predictive of Alzheimer's disease and impaired in schizophrenia and chronic drug users. An automated computer touchscreen PAL task for rats has been previously established. A pharmacologically validated PAL task for mice would be a highly valuable tool, which could be useful for a number of experimental aims including drug discovery. OBJECTIVES: This study sought to investigate the effects of systemic administration of cholinergic agents on task performance in C57Bl/6 mice. METHODS: Scopolamine hydrobromide (0.02, 0.2, and 2.0 mg/kg), dicyclomine hydrochloride (M(1) receptor antagonist; 2.0, 4.0, and 8.0 mg/kg), and donepezil hydrochloride (cholinesterase inhibitor; 0.03, 0.1, and 0.3 mg/kg) were administered post-acquisition in C57Bl/6 mice performing the PAL task. RESULTS: Scopolamine (0.2 and 2.0 mg/kg) and dicyclomine (at all administered doses) significantly impaired PAL performance. A significant facilitation in PAL was revealed in mice following donepezil administration (0.3 mg/kg). CONCLUSIONS: The present study shows that mice can acquire the rodent PAL task and that the cholinergic system is important for PAL task performance. M(1) receptors in particular are likely implicated in normal performance of PAL. The finding that mouse PAL can detect both impairments and improvements indicates that this task could prove to be a highly valuable tool for a number of experimental aims including drug discovery.

Pre-clinical and clinical pharmacology of selective muscarinic M3 receptor antagonists.

Muscarinic M3 receptor antagonists have therapeutic potential for the treatment of disorders associated with altered smooth muscle contractility or tone. These include irritable bowel syndrome (IBS), chronic obstructive airways disease (COAD) and urinary incontinence. Zamifenacin is a potent muscarinic receptor antagonist on the guinea pig ileum (pA2 value 9.27) with selectivity over M2 receptors in the atria (135-fold) and M1/M4 receptors in the rabbit vas deferens (78-fold). In addition, zamifenacin had lower affinity for the M3 receptor in the salivary gland (pKi 7.97). In animals, zamifenacin potently inhibited gut motility in the absence of cardiovascular effects and with selectivity over inhibition of salivary secretion. In healthy volunteers, zamifenacin inhibited small and large bowel motility and increased the rate of gastric emptying over a dose range which was associated with minimal anticholinergic side effects. These data show that zamifenacin, a selective muscarinic M3 receptor antagonist, was well tolerated in man and was efficacious as an inhibitor of gut motility. Further studies in patients are required with muscarinic M3 receptor antagonists to confirm efficacy against symptoms in diseases associated with altered smooth muscle contractility.