实验仪器使用技术

Neuronal Ca2+ signals occur in a very complex way. Direct imaging of Ca2+ changes in the soma, dendrites, and even single spines on fast time-scales greatly helps us understand the generation mechanism of diverse Ca2+ signaling events. However, Ca2+ imaging itself does not give information about ...

Calcium handling by mitochondria is important both because mitochondria can shape the cytosolic Ca2+ signals and because changes in mitochondrial Ca2+ concentration (M) are important for controlling physiological functions such as respiration or programmed cell death. Accur ...

A conventional borosilicate glass patch pipette is glued into a plastic jacket, forming the entity of a FlipTip�. One or two three-channel modules of recording tip sockets are mounted on a liquid handler platform to take up FlipTips. The tip sockets are connected to preamplifiers (HEKA) and to a suct ...

The aim of this chapter is to give a detailed, step-by-step description of a procedure for obtaining a batch of a desired peptide at the required level of purity. It is assumed that the initial synthesis will be done using an automated peptide synthesizer that performs solid-phase peptide synthesis ...

Quantification of mRNA levels can be performed using polymerase chain reaction (PCR)-based techniques (e.g., competitive reverse transcription PCR; RT-PCR), RNase protection, or Northern blotting. Northern blotting is less sensitive than the other techniques; however, the met ...

Automated electrophysiological assays are of great importance for modern drug discovery, and various approaches have been developed into practical devices. Here, we describe the automation of two-electrode voltage-clamp (TEVC) recording from Xenopus oocytes using the Roboo ...

In this study, we have demonstrated a method to organize cells in dissociated cultures using engineered chemical clues on the culture surface and determined their connectivity patterns. Although almost all elements of the synaptic transmission machinery between neurons or between ...

In order to explore the possibility of identifying toxins based on their effect on the shape of action potentials, we created a computer model of the action potential generation in NG108-15 cells (a neuroblastoma/glioma hybrid cell line). To generate the experimental data for model validat ...

Control of membrane voltage and membrane current measurements are of strong interest for the study of numerous aspects of skeletal muscle physiology and pathophysiology. The silicone-clamp technique makes use of a conventional patch-clamp apparatus to achieve whole-cell volta ...

This chapter describes methods to investigate mammalian cardiac channel properties at the single-channel level. Cell isolation is performed from adult heart by enzymatic digestion using the Langendorff apparatus. Isolation proceeding is suitable for rabbit, rat, and mouse hea ...

The method described here to differentiate mouse embryonic stem (ES) cells into cardiomyocytes is adapted from Maltsev et al. and results in a high percentage of spontaneously beating cardiomyocyte-like cells. In order to determine to what extent the differentiating ES cells resemble t ...

Even a complete understanding of the biophysical properties driving neuronal behavior would be insufficient to explain the interactions between neurons, neuronal assemblies, and brain regions. Exploring interactions between small numbers of synaptically coupled neuro ...

The cardiac long QT syndrome (LQTS) is characterized by a delayed repolarization of the ventricular myocytes, resulting in prolongation of the QT interval on the electrocardiogram and increased propensity to cardiac arrhythmias. Congenital LQTS has been linked to mutations in genes ...

Long-term potentiation is a synaptic mechanism thought to be involved in learning and memory. Long-term depression (LTD), an activity-dependent decrease in synaptic efficacy, may be an equally important mechanism that permits neural networks to store information more effective ...

The patch-clamp technique has allowed detailed studies on the electrical properties of neurons. Dye loading through patch pipettes has allowed characterizing the morphological properties of the neurons. In addition, the patch-clamp technique also allows harvesting mRNA from s ...

Mechanosensitive ion channels play an important role for the perception of mechanical signals such as touch, balance, or sound. Here, a new experimental strategy is presented providing well-defined access to single mechanosensitive ion channels in living cells. As a representative e ...

Single-channel recording provides molecular insights that are nearly unattainable from macroscopic measurements. Analysis of the data, however, has proven to be a difficult challenge. Early approach relies on the half-amplitude threshold detection to idealize the data into dwe ...

The Xenopus laevis oocyte is a widely used system for heterologous expression of exogenous ion channel proteins (1, 2). Among other advantages, these easy to obtain, mechanically and electrically stable, large-sized cells enable multiple types of electrophysiological recordings: ...

The functional properties of central synapses are difficult to study because they can be modulated either presynaptically or postsynaptically, each connection has multiple contacts and release at each contact is stochastic. Moreover, studying central synapses with electrop ...

The introduction of pseudopeptide bonds (amide bond surrogates) into the peptide backbone during synthesis is now a common technique in peptide chemistry (1). These pseudo-peptide bonds are introduced in order to satisfy criteria such as stability to enzymatic degradation, transi ...