Application of electrophysiological technique in toxicological study: From manual to automated patch-clamp recording

Highlights

• Patch-clamp technique is gold standard for ion channel studies.

• Manual patch-clamp recording has successfully been used in toxicological studies.

• Practical use of manual technique limited by low throughput and high skill requirement.

• High throughput screening promoted by evolving automated patch-clamp platform.

• Combination of manual and automated techniques is promising for toxicant screening.

Abstract

Ion channels are common targets of a range of environmental substances, including pharmaceuticals, toxins, metals and many organic pollutants. Patch-clamp technique is gold standard in studying ion channel characteristics, contributing to the improved accuracy in active compound identification, the increasing studies of molecular mechanism behind diseases, and the better prediction of potential toxicants. Nevertheless, low throughput and high technical capability requirement limit the application of conventional manual patch-clamp recording and an emerging transformation towards automation and simplification is rising. The evolving automated patch-clamp technique shows obvious advances over manual technique in many aspects, including reproducibility, throughput, operability and standardization. The application of emerging automated patch-clamp platform has created valuable consequences in many research areas, especially in high throughput screening. In this study, we provided an overview on the progresses made by conventional manual and emerging automated patch-clamp techniques in toxicological studies.

Introduction

Ion channels represent a membrane-bound protein family that regulates and facilitates ion movement across membranes. As a result of their special structure, many ion channels have ion selectivity, mainly consisting of sodium, potassium, calcium, and chlorine channels. Moreover, each type of ion channel includes several subtypes, representing different proteins associated with various drug or toxin targets. The physiological function of ion channels involves participation in cell activities such as cell differentiation, neural transmission, and cell migration and apoptosis. Therefore, abnormal ion channel activity triggers human diseases such as skeletal muscle disorders, cardiac arrhythmias, and epilepsy [[1], [2], [3]]. Indeed, ion channels can act as both a target of drugs to cure disease and as a target of toxic substances, leading to adverse reactions.

Previous studies have reported on the regulation of ion channels by environmental substances leading to various malfunctions [4,5]. For example, tetrodotoxin (TTX), a natural toxin found in many marine animals, is a powerful sodium channel inhibitor that can selectively block voltage-gated sodium channels in nerves and muscles, leading to the disruption of neuronal and muscular activities [6]. Drugs in clinical use have been shown to block the cardiac K_v11.1 voltage-gated potassium channel, with dangerous side effects, resulting in the prohibition of various drugs affecting this ion channel [2]. Similarly, certain environmental pollutants have been shown to prevent normal neural transmission based on their interaction with ion channels. Polychlorinated biphenyls (PCBs), a family of persistent organic pollutants, can interact with ryanodine receptors in neurons and change the spatial and temporal properties of calcium signals [7]. Thus, the toxicity of many chemicals found in the environment is based on their effect on ion channels, suggesting the necessity of further research on this topic.

At present, the patch-clamp technique has become the gold standard for studies on ion channel properties. Conventional manual patch-clamp records the channel current or voltage in real time and provides precise electrophysiological data with respect to ion channel function at a single-cell or single-channel level [8]. Recently emerging automated patch-clamp techniques are now able to provide a precise measurement of ion channel features with an improved throughput and a reduced skill requirement [9,10]. In many toxicological studies, automated patch-clamp has been used to provide adequate information on ion channel activities and greatly saves time and efforts. For instance, automated patch-clamp systems have been applied for the early safety assessment of a large number of drug candidates according to their potency to block the cardiac human ether-a-go-go-related gene (hERG) channel. These studies have led to valuable quantitative statistics, suggesting the potency of this evolving technique [2]. As the rapid and precise screening of toxicants and the identification of toxic effects are greatly needed in toxicology, an overview of automated patch-clamp systems and their potential to be applied in toxicological studies is of significance.

To the best of our knowledge, there are a limited number of reviews discussing the application of automated patch-clamp techniques in toxicological studies due to the techniques having just emerged in recent years. Herein, we first briefly describe the conventional manual and emerging automated patch-clamp techniques followed by a general overview of the application of manual patch-clamp technique in studying the effects of environmental substances on ion channels. Recent advances in toxicological studies resulting from the automation of the patch-clamp techniques are then discussed. Finally, we compare the manual and automated patch-clamp techniques on multiple dimensions, including cell preparation, throughput, operability, and data quality.

The literature was screened via Web of Science for articles published between January 1990 and March 2020, with key topic terms such as patch-clamp, high throughput electrophysiology, and ion channel, along with specific pollutants, metals, toxins, or drugs.