Inhibition of ligand binding to G protein-coupled receptors by arachidonic acid.

Arachidonic acid (AA), released in response to muscarinic acetylcholine receptor (mAChR) stimulation, previously has been reported to function as a reversible feedback inhibitor of the mAChR. To determine if the effects of AA on binding to the mAChR are subtype specific and whether AA inhibits ligand binding to other G protein-coupled receptors (GPCRs), the effects of AA on ligand binding to the mAChR subtypes (M1, M2, M3, M4, and M5) and to the micro-opioid receptor, beta2-adrenergic receptor (beta2-AR), 5-hydroxytryptamine receptor (5-HTR), and nicotinic receptors were examined. AA was found to inhibit ligand binding to all mAChR subtypes, to the beta2-AR, the 5-HTR, and to the micro-opioid receptor. However, AA does not inhibit ligand binding to the nicotinic receptor, even at high concentrations of AA. Thus, AA inhibits several types of GPCRs, with 50% inhibition occurring at 3-25 MuM, whereas the nicotinic receptor, a non-GPCR, remains unaffected. Further research is needed to determine the mechanism by which AA inhibits GPCR function.

The window of opportunity for treatment of focal cerebral ischemic damage with noninvasive intranasal insulin-like growth factor-I in rats.

Intracerebroventricular injection of insulin-like growth factor (IGF)-I has been shown to protect against stroke in rats. This method of delivery is not practical in human beings, as it requires an operation with risk of infection and other complications. Intranasal (i.n.) delivery offers a noninvasive method of bypassing the blood-brain barrier to deliver IGF-I to the brain. This study delineates the window of opportunity for treatment of focal cerebral ischemic damage using i.n. IGF-I after middle cerebral artery occlusion (MCAO). Rats were allowed to survive 7 days after 2 hours of MCAO. Infarct volume, apoptosis after 7 days, and neurologic deficit scores from the postural reflex and adhesive tape tests assessing motor-sensory and somatosensory functions, respectively, at 1 to 7 days were used to evaluate the efficacy of i.n. IGF-I (150 microg) administered at different times after MCAO. I.n. IGF-I significantly reduced infarct volume by 54%; and 39%; versus control when administered at 2 or 4 hours, respectively, after the onset of MCAO (P < .05) and improved motor-sensory and somatosensory functions (P < .05) when administered 2 hours after the onset of MCAO. In addition, treatment with i.n. IGF-I at 2, 4, or 6 hours after MCAO decreased apoptotic cell counts by more than 90%; in the hemisphere ipsilateral to the occlusion. I.n. IGF-I is a promising treatment for stroke with a therapeutic window of opportunity for up to 6 hours after the onset of ischemia. This noninvasive method provides a simpler, safer, and potentially more cost-effective method of delivery than other methods currently in use.

Inactivation of the human brain muscarinic acetylcholine receptor by oxidative damage catalyzed by a low molecular weight endogenous inhibitor from Alzheimer's brain is prevented by pyrophosphate analogs, bioflavonoids and other antioxidants.

Oxidative stress has been implicated as a contributing factor to neurodegeneration in Alzheimer's disease. An endogenous, low molecular weight (LMW) inhibitor from Alzheimer's brain inactivates the human brain muscarinic acetylcholine receptor (mAChR). The inhibitor prevents agonist and antagonist binding to the mAChR as assessed by radioligand binding studies. The LMW endogenous inhibitor, which has components with molecular weights between 100 and 1000 Da, requires dissolved oxygen and glutathione. Prevention of inactivation of the mAChR with peroxidase suggests that the LMW endogenous inhibitor generates peroxide. Heme, previously shown to be present in the LMW endogenous inhibitor, also inactivates the mAChR in the presence of peroxide. Free radical damage to the muscarinic receptor by the endogenous inhibitor can be prevented through the use of naturally occurring antioxidants including bilirubin, biliverdin, carnosol, myricetin and quericetin. In addition, pyrophosphate, imidodiphosphate, bisphosphonates and related compounds also protect the muscarinic receptor from free radical damage. Inactivation of the mAChR by the LMW endogenous inhibitor is likely to be a factor in the continual decline of Alzheimer's patients, even those taking acetylcholinesterase inhibitors. Natural antioxidants and pyrophosphate analogs may improve the effectiveness of acetylcholinesterase inhibitors and prove useful in the treatment and prevention of Alzheimer's disease since the muscarinic acetylcholine receptor is required for memory, and decreased cholinergic function is a critical deficit in Alzheimer's disease.

Delivery of Nerve Growth Factor to the Brain via the Olfactory Pathway.

Purpose: To assess the potential of delivering nerve growth factor (NGF) to the brain along the olfactory neural pathway for the treatment of Alzheimer's disease. Methods: Recombinant human NGF (rhNGF) was given as nose drops to anesthetized rats. The rhNGF concentrations in the brain were determined by enzyme-linked immunosorbent assay (ELISA). Results: Following olfactory administration, rhNGF reached the brain within an hour, achieving a concentration of 3400 pM in the olfactory bulb, 660­2200 pM in other brain regions and, 240 pM and 180 pM in the hippocampus and the amygdala, respectively. In contrast, little or no rhNGF was found in the brain following intravenous administration. Conclusions: A significant amount of rhNGF can be delivered to the brain via the olfactory pathway. The detection of rhNGF by ELISA indicates that rhNGF is delivered to the brain relatively intact. The rapid appearance of rhNGF in the brain suggests that it may be transported by an extraneuronal route into the brain via intercellular clefts in the olfactory epithelium. Further work to clarify the transport mechanism is underway. The olfactory pathway is a promising, non-invasive route for drug delivery to the brain, which has potential for the treatment of neurodegenerative diseases including Alzheimer's disease.