The N-substituted 3␣-[bis(4Ј-fluorophenyl)methoxy]tropanes (AHN 2-003, AHN 1-055, AHN 2-005, and JHW 007) bind with high affinity to the dopamine transporter and inhibit dopamine uptake more potently than cocaine, but they demonstrate behavioral profiles in animal models of psychostimulant abuse that are unlike that of cocaine. The objective of this study was to characterize the in vitro permeability, brain distribution, and pharmacokinetics of the benztropine (BZT) analogs. Transport studies of cocaine and the BZT analogs (10 Ϫ4 M) were conducted across bovine brain microvessel endothelial cells. Male Sprague-Dawley rats (ϳ300 g) were administered BZT analogs (10 mg/kg) or cocaine (5 mg/kg) via the tail vein. Blood and brain samples were collected over 36 h and assayed using UV-high-performance liquid chromatography. Transport of both AHN 1-055 (2.15 ϫ 10 Ϫ4 cm/s) and JHW 007 (2.83 ϫ 10 Ϫ4 cm/s) was higher (p Ͻ 0.05) than that of cocaine (1.63 ϫ 10 Ϫ4

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Previous studies have demonstrated that chronic mechanical strain produced by continuous positive airway pressure (CPAP) reduces in vivo airway reactivity in rabbits and ferrets. For CPAP to potentially have a therapeutic benefit for asthmatic subjects, the reduction in airway responsiveness would need to persist for 12-24 h after its discontinuation, require application for only part of the day, and be effective in the presence of atopic airway inflammation. In the present study, airway responsiveness to acetylcholine or methacholine was measured during mechanical ventilation following three different protocols in which active, nonanesthetized, tracheotomized rabbits were treated with High vs. Low CPAP (6 vs. 0 cmH(2)O). 1) High CPAP was applied continuously for 4 days followed by 1 day of Low CPAP; 2) High CPAP was applied at night and Low CPAP during the daytime for 4 days, and 3) High CPAP was applied for 4 days in animals following ovalbumin (Ova) sensitization and challenge. For all three protocols, treatment with High CPAP resulted in significantly reduced airway responsiveness compared with treatment with Low CPAP. Cumulatively, our in vivo results in rabbits suggest that high CPAP, even when applied only at night, produces a persistent reduction of airway responsiveness. In addition, CPAP reduces airway responsiveness even in the presence of atopic airway inflammation.

Multidrug resistance (MDR), mediated by P-glycoprotein (Pgp) has been identified as altering the disposition of structurally diverse compounds. Previous in vitro studies in bovine brain microvascular endothelial cells and MCF/Adr [Adriamycin (doxorubicin)-resistant human breast cancer] cells displayed that the transport of enaminone anticonvulsants was influenced by Pgp. Therefore the objectives of this study was to further evaluate the influence of Pgp on the pharmacokinetics and tissue distribution of the enaminone analogs. mdr1ab (ϩ/ϩ) and mdr1ab (Ϫ/Ϫ) male mice (20 Ϯ 5 g) were administered DM5 (methyl 4-[(4Ј-chlorophenyl)amino]-6-methyl-2-oxo-3-cyclohexene-1-carboxylate) or DM44 (12.5 mg/kg, i.v.). Cohorts (n ϭ 3) were sacrificed over a 12-h period, and samples were analyzed by a validated UV-high performance liquid chromatography assay method. Population analysis was used to estimate pharmacokinetic parameters and partition coefficients were determined for tissues. The clearance (0.51 versus 0.33 l/h/kg) and V d (1.25 versus 0.93 l/kg) of DM5 were found to be higher (p Ͻ 0.05), however the area under the curve (26.1 versus 38.2 g/ml ⅐ h) was lower (p Ͻ 0.05) in mdr1a/1b (Ϫ/Ϫ) versus mdr1a/1b (ϩ/ϩ) mice, respectively. Similar findings were observed for DM44. Tissues known to express Pgp such as the heart, liver, lung, and brain displayed 2-fold or higher tissue levels in mdr1a/1b (Ϫ/Ϫ) versus mdr1a/1b (ϩ/ϩ) mice. These results strongly suggest that Pgp may influence enaminone tissue distribution and pharmacokinetics and may play a significant role in the effective treatment of epilepsy with these analogs.

The p21-activated protein kinases (Paks) have been implicated in the regulation of smooth muscle contractility, but the physiologic effects of Pak activation on airway reactivity in vivo are unknown. A mouse model with a genetic deletion of Pak1 (Pak1 −/−) was used to determine the role of Pak in the response of the airways in vivo to challenge with inhaled or intravenous acetylcholine (ACh). Pulmonary resistance was measured in anesthetized mechanically ventilated Pak1 −/− and wild type mice. Pak1 −/− mice exhibited lower airway reactivity to ACh compared with wild type mice. Tracheal segments dissected from Pak1 −/− mice and studied in vitro also exhibited reduced responsiveness to ACh compared with tracheas from wild type mice. Morphometric assessment and pulmonary function analysis revealed no differences in the structure of the airways or lung parenchyma, suggesting that that the reduced airway responsiveness did not result from structural abnormalities in the lungs or airways due to Pak1 deletion. Inhalation of the small molecule synthetic Pak1 inhibitor, IPA3, also significantly reduced in vivo airway responsiveness to ACh and 5-hydroxytryptamine (5-Ht) in wild type mice. IPA3 inhibited the contractility of isolated human bronchial tissues to ACh, confirming that this inhibitor is also effective in human airway smooth muscle tissue. The results demonstrate that Pak is a critical component of the contractile activation process in airway smooth muscle, and suggest that Pak inhibition could provide a novel strategy for reducing airway hyperresponsiveness.