Novel protocol for multiple-dose oral administration of the L-type Ca2+ channel blocker isradipine in mice: A dose-finding pharmacokinetic study

Studies in genetically modified animals and human genetics have recently provided new insight into the role of voltage-gated L-type Ca2+ channels in human disease. Therefore, the inhibition of L-type Ca2+ channels in vivo in wildtype and mutant mice by potent dihydropyridine (DHP) Ca2+ channel blockers serves as an important pharmacological tool. These drugs have a short plasma half-life in humans and especially in rodents and show high first-pass metabolism upon oral application. In the vast majority of in vivo studies, they have therefore been delivered through parenteral routes, mostly subcutaneously or intraperitoneally. High peak plasma concentrations of DHPs cause side effects, evident as DHP-induced aversive behaviors confounding the interpretation of behavioral readouts. Nevertheless, pharmacokinetic data measuring the exposure achieved with these applications are sparse. Moreover, parenteral injections require animal handling and can be associated with pain, discomfort and stress which could influence a variety of physiological processes, behavioral and other functional readouts. Here, we describe a noninvasive oral application of the DHP isradipine by training mice to quickly consume small volumes of flavored yogurt that can serve as drug vehicle. This procedure does not require animal handling, allows repeated drug application over several days and reproducibly achieves peak plasma concentrations over a wide range previously shown to be well-tolerated in humans. This protocol should facilitate ongoing nonclinical studies in mice exploring new indications for DHP Ca2+ channel blockers.

近年来，基于转基因动物与人类遗传学的研究，为电压门控L型钙通道（voltage-gated L-type Ca²⁺ channels）在人类疾病中的作用机制提供了全新视角。因此，采用强效二氢吡啶类（dihydropyridine, DHP）钙通道阻滞剂，在野生型及突变小鼠体内抑制L型钙通道，成为一项关键的药理学研究工具。此类药物在人类尤其是啮齿类动物体内的血浆半衰期较短，口服给药时会发生显著的首过代谢（first-pass metabolism）。因此，绝大多数在体研究均通过胃肠外给药途径（parenteral routes）给予此类药物，以皮下注射（subcutaneously）或腹腔注射（intraperitoneally）最为常见。二氢吡啶类药物的血浆峰浓度过高时会引发不良反应，表现为药物诱导的厌恶行为（aversive behaviors），这会干扰行为学检测结果（behavioral readouts）的解读。尽管如此，针对此类给药方式所实现的药物暴露量的药代动力学数据（pharmacokinetic data）仍较为稀缺。此外，胃肠外注射需要对动物进行操作，可能引发疼痛、不适与应激反应，进而影响多种生理过程、行为学及其他功能检测结果。本文中，我们介绍一种无创口服给药方案（noninvasive oral application）：通过训练小鼠快速摄取少量调味酸奶（flavored yogurt）作为药物载体（drug vehicle），给予二氢吡啶类药物伊拉地平（isradipine）。该操作无需对动物进行固定处理，可在数日内重复给药，且可稳定获得在人体中已被证实具有良好耐受性的宽范围血浆峰浓度。本实验方案将助力当前以小鼠为模型的非临床研究，进一步探索二氢吡啶类钙通道阻滞剂的新适应症。