神经生物学

Calcium ions are the most ubiquitous and pluripotent signalling molecules, which regulate a wide array of physiological and pathological reactions. The specific system, controlling cellular Ca2+ homeostasis appeared very early in the evolution, being initially survival syst ...

Glial cells are besides neurons the second major cell type of nervous systems and are either of neuroectodermal (macroglia) or mesodermal (microglia) origin. As electrically non-excitable cells, they employ calcium signals in response to most external stimuli, which initiate cellu ...

This chapter overviews the use of acetoxymethyl (AM) ester-based multi cell bolus loading (MCBL) technique for in vivo Ca2+ imaging of neural circuits. This technique provides anatomically targeted, rapid and non-invasive staining of both neurones and glia with small molecule Ca2+ indi ...

The intracellular calcium concentration is one key parameter triggering numerous intracellular signalling pathways in neuronal cells. The development of optical techniques like fast confocal or 2-photon microscopy has made it possible to measure calcium dynamics even in sub- ...

In the last two decades, the study of Ca2+ homeostasis in living cells received a great impulse by the explosive development of genetically encoded Ca2+-indicators. The cloning of the Ca2+-sensitive photoprotein aequorin and of the green fluorescent protein (GFP) from the jellyfish Aequo ...

In this chapter, we review the theoretical and experimental foundations underling a quantitative approach to Ca2+ imaging, discuss equilibrium conditions and their violations and present a computational framework that can be used to estimate the spatial and temporal dynamics of Ca2+ ...

Ca2+-sensitive microelectrodes are time-consuming to make and require large robust cells. But, they do not add to buffering and do not require expensive equipment. I describe how to make and use the electrodes and briefly consider the leakage problem.

Calcium indicators are widely used to monitor activity in living neuronal tissue because of the tight relation between action potential firing and increases in the intracellular calcium concentration. Here, we describe the use of genetically encoded calcium indicators (GECIs) of t ...

The endoplasmic reticulum (ER) is a complex and highly dynamic three-dimensional intracellular membranous system, which acts as a dynamic calcium store in the majority of eukaryotic cells. The special arrangement of intra-ER Ca2+ buffers, characterized by low affinity for Ca2+, in comb ...

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 ...