Intracellular environment, membranes

Ion Channels. The excitable cell maintains an asymmetric distribution across both the plasma membrane, defining the extracellular and intracellular environments, as well as the intracellular membranes which define the cellular organelles. This maintained a symmetric distribution of ions serves two principal objectives. It contributes to the generation and maintenance of a potential gradient and the subsequent generation of electrical currents following appropriate stimulation. Moreover, it permits the ions themselves to serve as cellular messengers to link membrane excitation and cellular... 

In addition to their function as a permeability barrier to the extracellular environment, membranes also fulfil important tasks inside most eukaryotic cells and in some bacteria. One crucial role is the separation of different cell compartments. A few examples of intracellular membranes may illustrate the large variety of membrane functions ... 

The plasma membrane forms a boundary between the extra- and intracellular environments whereas membranes within a cell form boundaries between the organelles and the cytosol. These are discussed in other chapters, whereas the material in this chapter focuses on the plasma membrane. A primary function of this membrane is to serve as a barrier to prevent the entry of some molecules and ions into the cell and to retain others within the cell (Table 5.1). The plasma membrane has other roles, which are related to the presence of proteins within or attached to the membrane. These are ... 

Since aminoglycoside complexes with membrane phosphoinositides are extremely stable, they could well exist in an intracellular environment with concentrations of aminoglycosides that are achieved in chemotherapeutic treatment. 

These properties are likely to have an important influence on the behavior of intact biochemical systems, e.g., within the living cell, enzymes do not function in dilute homogeneous conditions isolated from one another. The postulates of the Michaelis-Menten formalism are violated in these processes and other formalisms must be considered for the analysis of kinetics in situ. The intracellular environment is very heterogeneous indeed. Many enzymes are now known to be localized within 2-dimensional membranes or quasi 1-dimensional channels, and studies of enzyme organization in situ [26] have shown that essentially all enzymes are found in highly organized states. The mechanisms are more complex, but they are still composed of elementary steps governed by fractal kinetics. 

Spicule formation takes place within the intracellular environment. The spicules, however, are much larger than individual cells. This is achieved by many cells fusing their membranes to enclose an extended space (called a syncytium) (Fig. 1.9). Spicule formation takes place inside a membrane delineated vacuole within this space [70, 71]. The size and the shape of the syncytium constantly increase and change during growth of the spicule [72, 73]. In fact, a freeze-fracture TEM study of the relation between the membrane and the growing spicule shows that the membrane is always juxtaposed to the spicule surface. There is thus no bulk solution within which the spicule forms [74], Thus spicule... 

Reduction of the toxin and translocation of the L chain into the cytoplasm To gain access to the cytoplasm, the L chain needs to cross the membrane of the endocytic compartment. For this translocation, the H chain is required, probably by forming a proteinaceous translocation complex in the membrane that exposes (and possibly releases) the L chain to the cytoplasm. In the reductive intracellular environment, the disulfide bond linking the H and L chains is reduced (Kistner and Habermann, 1992). 

Monitoring of the intracellular redox activity in eukaryotic cells imposes the requirement that the utilized mediator is capable of readily crossing the plasma membrane into the intracellular environment to communicate with the enzyme(s), the activity of which is to be monitored. This strictly requires the utilization of a lipophilic mediator that can diffuse through the plasma membrane. Using chip based amperometric detection on S. cerevisiae, menadione was shown to possess the desired properties [28]. Figure 3 depicts the functional principle of the chip based detection technique to monitor CRE in eukaryotic cells, which aside from the lipophilic menadione,... 

Integral proteins are embedded in the membrane itself, to some degree, and are referred to as transmembrane proteins if they extend from one side of the membrane to the other. Typically these proteins have multiple domains or regions that are either primarily hydrophobic, if embedded in the lipid bilayer, or hydrophilic if localized in the extra- or intracellular environment. More complicated tertiary and quaternary protein domain structures allow for the formation of channels or pores where appropriate arrangement of hydrophilic and hydrophobic amino acids on the internal surface of each channel or pore dictates which molecules may enter or bind for subsequent translocation from one side of the membrane to the other. Based on this, some proteins exhibit considerable substrate specificity (e.g., GLUTl a glucose transporter Scheepers et al., 2004) whereas others appear much less specific (e.g., P-glycoprotein Leslie et al., 2005). 

In multicellular organisms, thin lipid membranes serve as semipermeable barriers between aqueous compartments (Figure 5.1). The plasma membrane of the cell separates the cytoplasm from the extracellular space endothelial cell membranes separate the blood within the vascular space from the rest of the tissue. Properties of the lipid membrane are critically important in regulating the movement of molecules between these aqueous spaces. While certain barrier properties of membranes can be attributed to the lipid components, accessory molecules within the cell membrane—particularly transport proteins and ion channels—control the rate of permeation of many solutes. Transport proteins permit the cell to regulate the composition of its intracellular environment in response to extracellular conditions. 

Lipid bilayers are impermeable to ions and other charged species (Figure 5.6), but ions such as and K are found in abundance in both the extracellular and intracellular environment. Active transport proteins can move ions across membranes, even moving molecules against a concentration... 

The development of in vitro brain slice and isolated neuron techniques has greatly facilitated detailed studies of the electrophysiology of a wide range of neuronal types in the adult and neonatal vertebrate central nervous system (CNS). Particularly advantageous are the greater mechanical stability that these preparations provide over in vivo models and the control allowed over the composition of the extracellular environment. In addition, the development of the patch-clamp technique has opened up the possibility of direct access to the intracellular environment via internal patch pipet solutions. In combination, these approaches have enabled detailed investigations of neuronal membrane properties, the cellular actions of neurotransmitters, and synaptic mechanisms. 

Knowing that damaging the integrity of cells impaired the a-oxidation process, a search for possible cofactors was initiated in rat hepatocytes permeabilized with Staphylococcus aureus toxin. In such systems, the intracellular environment can be varied experimentally but the integrity of the intracellular membranes is conserved. 

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