神经生物学

It is generally agreed that ion channels are operationally composed of two functional domains: The selectivity filter comprises the part of the protein that determines which ion may pass and which is retained; the gate determines under which conditions the selected ions may pass. Named acco ...

Muscle contraction is affected by the rapid release of Ca2+ ions through Ca2+- conducting channels localized in an intracellular mem- brane compartment, the sarcoplasmic reticulum (SR). The rabbit skel- etal muscle SR Ca2+ release channel has been identified as a high-affinity receptor for ...

Calcium channels are an essential part of the cellular signal trans duction system, since they produce changes in cytosolic calcium. Three types of voltage-dependent calcium channels (T, L, and N channels) have been identified by electrophysiological and pharmacological tech- niqu ...

Purification of receptor proteins has always been an important and challenging task in advancing the structural characterization and also in providing sequence data for the molecular cloning of the recep- tors. Some of the difficulties in the purification of receptors are:

A variety of biochemical and physiological studies in recent years have made it clear that the brain possesses specific receptors and puta- tive signal transduction pathways for insulin and the insulin-like growth factors (IGF-I and IGF-II). These observations have challenged tradi- t ...

This chapter is designed to serve as a guide for neurobiologists interested in studying neurotransmitter receptors and ion channels expressed in the oocytes of the African clawed frog Xenopls laevis. The emphasis is on routinely employed methods, based mostly on the experience gained in o ...

The typical mammalian genome contains 3 � lo9bp of DNA. This amount of DNA is believed to include 3-4 � lo4 unique protein- encoding genes. Even complex, highly specialized cells, such as neurons, express a small fraction, perhaps 20%, of all the possible genes, how- ever. A major problem in neurobiology, th ...

Recent progress in the field of molecular biology has provided a new method to localize an mRNA signal within a cell or tissue section. This technique, called “in situ hybridization histochemistry,” gives us dynamic information about gene expression at individual cellular lev- els. For the de ...

Because of the vast organizational complexity of the brain, it is crucial to study gene regulation at the level of individual cells. Most brain regions or individual neurons lying adjacent to each other may contain different neurotransmitters or different receptors, which may differen ...

Human mitochondrial DNA (mtDNA) is a closed circular genome of 16,569 bp (1), encoding 13 subunits of four enzyme complexes (Complexes I, III, IV, and V) in the oxidative phosphorylation system (2). Mutations of mtDNA have been demonstrated to be associated with various neuromuscular diseases. ...

Chemically synthesized oligonucleotides are powerful tools in the molecular biologist’s repertoire. This chapter describes the design of oligonucleotides and their applications, with particular reference to their use in the isolation and characterization of recombina ...

The polymerase chain reaction (PCR; 1) employs a pair of oligonucleotide primers, one sense and one antisense, with respect to the template DNA. During repeated cycles of heat denaturation, annealing of the oligonucleotide primers to the template and extension of the primers by the enzyme taq p ...

All cells have a large number of proteins; for instance, a single hepatocyte has about lo4 different proteins, and a number of them are involved in tissue-specific functions. In neuronal cells, there are additional 104 protein species, which are predicted, from kinetic of DNA-RNA hybridizati ...

The complexity of neural gene expression results from an inter- play of a multitude of signal transduction pathways and trans-acting transcription factors interacting at cis-acting DNA elements (1). Clearly, a major challenge is to define the pathways and molecular mechanism(s) inter ...

The appearance and maintenance of differentiated neural cell types in eukaryotic organisms can be largely attributed to characteristic spa- tial and temporal regulatory cascades evoking cell-specific gene expres- sion. Since gene expression causes information passage from D ...

This chapter describes procedures for purification of μ-calpain, m-calpain, and calpastatin simultaneously from a single tissue sample. The procedures and reagents described apply equally well to purification of any one or two of these three proteins from an appropriate tissue with a c ...

Although in recent years the molecular structure and function of calpains have been extensively investigated, some aspects of the roles and properties of these proteinases remain obscure, particularly their precise function in the cell, and the mechanisms inducing and promoting th ...

Determination of calpastatin levels in cells or tissues by Western blotting using antibodies, or by assay of calpain inhibitory activities in vitro, may be required to assess the role and the level of calpastatin, with respect to the levels of other components of the calpain system, in certain cell ...

Calpastatin is an endogenous inhibitor protein specific for calpain (1,2). Other calpain-inhibiting reagents such as EDTA, E-64, leupeptin, and calpain inhibitor I (N-acetyl-L-leucyl-L-leucyl-L-norleucinal), are frequently used in attempts to assess the role of calpain in degra ...

In many cases separation of calpains and calpastatin has to be performed with a relatively small amount of tissue, so that large-scale and complicated multiple-column procedures are not appropriate. In designing a method suitable for small amounts of tissue, it is important that the extrac ...