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The endocannabinoid system (ECS) plays a diverse role in human physiology ranging from the regulation of mood and appetite to immune modulation and the response to pain. Drug development that targets the cannabinoid receptors (CB 1 and CB 2 ) has been explored; however, success in the clinic has been limited by the psychoactive side effects associated with modulation of the neuronally expressed CB 1 that are enriched in the CNS. CB 2 , however, are expressed in peripheral tissues, primarily in immune cells, and thus development of CB 2 -selective drugs holds the potential to modulate pain among other indications without eliciting anxiety and other undesirable side effects associated with CB 1 activation. As part of a collaborative effort among industry and academic laboratories, we performed a high-throughput screen designed to discover selective agonists or positive allosteric modulators (PAMs) of CB 2 . Although no CB 2 PAMs were identified, 167 CB 2 agonists were discovered here, and further characterization of four select compounds revealed two with high selectivity for CB 2 versus CB 1 . These results broaden drug discovery efforts aimed at the ECS and may lead to the development of novel therapies for immune modulation and pain management with improved side effect profiles.

Over the past decade, high throughput screening (HTS) has become the focal point for discovery programs within the pharmaceutical industry. The role of this discipline has been and remains the rapid and efficient identification of lead chemical matter within chemical libraries for therapeutics development. Recent advances in molecular and computational biology, i.e., genomic sequencing and bioinformatics, have resulted in the announcement of publication of the first draft of the human genome. While much work remains before a complete and accurate genomic map will be available, there can be no doubt that the number of potential therapeutic intervention points will increase dramatically, thereby increasing the workload of early discovery groups. One current drug discovery paradigm integrates genomics, protein biosciences and HTS in establishing what the authors refer to as the "gene-to-screen" process. Adoption of the "gene-to-screen" paradigm results in a dramatic increase in the efficiency of the process of converting a novel gene coding for a putative enzymatic or receptor function into a robust and pharmacologically relevant high throughput screen. This article details aspects of the identification of lead chemical matter from HTS. Topics discussed include portfolio composition (molecular targets amenable to small molecule drug discovery), screening file content, assay formats and plating densities, and the impact of instrumentation on the ability of HTS to identify lead chemical matter.

Alzheimer's disease, in its early onset familial form, is known to be a heterogeneous disorder. This suggests that the different degenerative mechanisms, initiated by different genetic causes and ending in the shared phenotype of the disease, should intersect at some point in the degenerative cascade to form a 'bottleneck' from which the pathological features that are common to each of the genetic forms emerge. A growing body of evidence suggests that disturbances of energy metabolism may play a fundamental role in the onset and progression of Alzheimer's disease. In light of this, we propose a 'mitochondrial bottleneck hypothesis', which unifies the various forms of the disease in which different causes lead to the disorder via disturbances of mitochondrial function. The characterization of such a bottleneck would present a unique target for therapeutic intervention because it would be the earliest point in a neurodegenerative cascade shared by all forms of Alzheimer's disease, independent of cause.

We describe a novel compound, (-)-N-(2-ethoxyphenyl)-N'-(1,2,3-trimethylpropyl)-2-nitroethene-1, 1-diamine), Bay x9227, that demonstrates dose-dependent hyperpolarizing activity of remarkable potency (EC50 3 picomolar) and selectivity for CNS neurons and clonal neurotypic cells compared to smooth muscle cells. Single cell membrane potential measurements were obtained in physiologic buffer using the fluorescent probe, bisoxonol. Unlike K+ATP-channel activators including its (+)-enantiomer (Hoffman et al., 1993, Biochem. Biophys. Res. Commun. 190(2), 551), this activity was insensitive to glibenclamide antagonism. These data suggest a novel pharmacologic site for effecting neuroselective hyperpolarization.

Although cocaine has a fascinating and complex medicinal history in man, its natural function in plants is unknown. The present studies demonstrate that cocaine exerts insecticidal effects at concentrations which occur naturally in coca leaves. Unlike its known action on dopamine reuptake in mammals, cocaine's pesticidal effects are shown to result from a potentiation of insect octopaminergic neurotransmission. Amine-reuptake blockers of other structural classes also exert pesticidal activity with a rank order of potency distinct from that known to affect vertebrate amine transporters. These findings suggest that cocaine functions in plants as a natural insecticide and that octopamine transporters may be useful sites for targeting pesticides with selectivity toward invertebrates.

Background and Purpose: The causes of volume depletion and hyponatremia after subarachnoid hemorrhage are not fully understood but may be in part due to natriuresis or "cerebral salt wasting." Because previous studies using infrequent hormone sampling have given inconsistent results, we determined if elevations in atrial natriuretic factor concentrations preceded negative sodium and fluid balances., Methods: We measured diurnal atrial natriuretic factor and vasopressin concentrations and sodium balance for 5 days in 14 consecutive patients after aneurysmal subarachnoid hemorrhage., Results: Plasma concentrations of atrial natriuretic factor on admission were elevated in subarachnoid hemorrhage patients (mean +/- SD 106 +/- 59 pg/ml) compared with acutely ill controls (39 +/- 30 pg/ml). In eight patients, high peak concentrations of atrial natriuretic factor, greater than 300 pg/ml or a twofold increase above baseline, were followed by natriuresis and a negative sodium balance. Three patients, two of whom became hyponatremic, developed cerebral infarcts after natriuresis. Vasopressin concentrations were slightly elevated just after hemorrhage but subsequently declined to normal values., Conclusions: A markedly increased atrial natriuretic factor concentration precedes natriuresis in some patients and, with other abnormalities of water handling possibly including a relatively diminished vasopressin concentration, may cause volume depletion. Patients with natriuresis appear to be at increased risk for delayed cerebral infarction after subarachnoid hemorrhage.

An octopamine receptor photoaffinity probe was used to label membranes from the light organs of Photinus pyralis, a tissue highly enriched in octopamine receptors. Labeling was concentrated in a glycoprotein of 75 +/- 2 kDa with lesser labeling of a 79 +/- 2 kDa component. Labeling could be displaced by prior incubation with octopamine, mianserin, cyproheptadine, phentolamine or propranolol, with a relative potency that correlated with the ability of these same agents to modulate light organ octopamine-sensitive adenylate cyclase. The 75 kDa binding protein was isolated and its N-terminal amino acid sequence was determined.

Octopamine, a putative phenylethylamine neurotransmitter present in the firefly, has been found to be a potent stimulator of cyclic AMP synthesis in the larval light organ. In the same tissue, octopamine causes a small inhibition of phosphodiesterase activity. Because of the relatively simple anatomical relationships present in the larval light organ, compared with that of the adult, this preparation should offer an attractive model for studying the biochemistry of neurohumoral control of light emission.

Recent studies indicate that selective beta 2-adrenoceptor antagonists may be of use in the treatment of glaucoma. ICI 118,551, its desmethyl-, didesmethyl-, and ethyl- analogues, and a putative highly beta 2-selective binary aryloxypropanolamine (4d) have been evaluated for their ability to inhibit beta-adrenergic-stimulated adenylate cyclase activity in rabbit ciliary process and heart. Potency ratios (Ki heart/Ki ciliary process) were 440 for ICI 118,551; 9.3 for the desmethyl analogue; 8.2 for the didesmethyl analogue; 720 for the ethyl analogue; and 11 for the binary aryloxypropanolamine. The values for the ethyl derivative of ICI 118,551 indicate that it is the most oculoselective beta-adrenoceptor antagonist yet reported.