classification of ion channels Ion channels are classified according to their electrophysiological properties, drug sensitivity, and by molecular cloning Ligand (agonist)-gated ion channels(direct-coupled; G-protein-coupled; sec- ond messenger-coupled), which include acetylcholine receptors(muscle(nico- tinic); neural), glycine receptors, GABA A receptors and glutamate receptors. Calcium channels are present in cell membranes in smooth muscle, cardiac muscle and other tissues, and in cellular organelle membranes such as the sarcoplasmic reticulum and mitochondria Calcium functions as a primary generator of the cardiac action potential and as an intracellular second messen- ger. Calcium channels are further subdivided into three subgroups based on their threshold for activation and on the spread of inactivation: L-type (long-lasting): slowly inactivating: high threshold calcium conduc- tance;sensitive to dihydropyridines; involved in excitation-contraction coupling in smooth and cardiac muscle (where they carry current in the plateau phase of the action potential), and in excitation-secretion coupling in endocrine cells and in some neurons T-type(transient): low voltage activated, rapidly inactivated N-type(neuronal): transient, high threshold calcium conductance; blocked by The T and L channels are located in smooth and cardiac muscle tissue, whereas the n channels are located only in neuronal tissue Calcium channel blockers interact with the L-type calcium channel and onsist of four classes of drugs: the 1, 4-dihydropyridine derivatives(nifedipine, nimodipine, amlodipine), the phenylalkyl-amines(verapamil), the benzothiaze pines(diltiazem), and a diarylaminopropylamine ether( bepridil) Potassium channels are tetrameric and composed of four identical peptide subunits(alpha subunits) that are symmetrically arranged to form a conical pore that spans the cell membrane. Many potassium channels also contain auxiliary that may alter electrophy properties, expression levels or expression patterns. They are divided into Six transmembrane-helix voltage-gated channels, which are activated by membrane depolarisatio Two transmembrane-span G-protein-coupled inward rectifying channels which favour the influx rather than efflux of potassium ions. Calcium-activated channels, which are sensitive to intracellular calcium concentrations. Large conductance: blocked by charybdotoxin and iberiotoxin Small conductance: blocked by apamin
Classification of ion channels Ion channels are classified according to their electrophysiological properties, drug sensitivity, and by molecular cloning. * Ligand (agonist)-gated ion channels (direct-coupled; G-protein-coupled; second messenger-coupled), which include acetylcholine receptors (muscle (nicotinic); neural), glycine receptors, GABA A receptors and glutamate receptors. * Calcium channels are present in cell membranes in smooth muscle, cardiac muscle and other tissues, and in cellular organelle membranes such as the sarcoplasmic reticulum and mitochondria. Calcium functions as a primary generator of the cardiac action potential and as an intracellular second messenger. Calcium channels are further subdivided into three subgroups based on their threshold for activation and on the spread of inactivation: L-type (long-lasting): slowly inactivating; high threshold calcium conductance; sensitive to dihydropyridines; involved in excitation–contraction coupling in smooth and cardiac muscle (where they carry current in the plateau phase of the action potential), and in excitation-secretion coupling in endocrine cells and in some neurons. T-type (transient): low voltage activated, rapidly inactivated. N-type (neuronal): transient, high threshold calcium conductance; blocked by omega-conotoxin The T and L channels are located in smooth and cardiac muscle tissue, whereas the N channels are located only in neuronal tissue. Calcium channel blockers interact with the L-type calcium channel and consist of four classes of drugs: the 1,4-dihydropyridine derivatives (nifedipine, nimodipine, amlodipine), the phenylalkyl-amines (verapamil), the benzothiazepines (diltiazem), and a diarylaminopropylamine ether (bepridil). * Potassium channels are tetrameric and composed of four identical peptide subunits (alpha subunits) that are symmetrically arranged to form a conical pore that spans the cell membrane. Many potassium channels also contain auxiliary proteins, beta subunits, that may alter electrophysiological or biophysical properties, expression levels or expression patterns. They are divided into: Six transmembrane-helix voltage-gated channels, which are activated by membrane depolarisation. Two transmembrane-span G-protein-coupled inward rectifying channels, which favour the influx rather than efflux of potassium ions. Calcium-activated channels, which are sensitive to intracellular calcium concentrations: Large conductance: blocked by charybdotoxin and iberiotoxin; Small conductance: blocked by apamin. Cell physiology 10
Leak channels, with an apparent lack of gating control. Sodium channels are discrete, four domain, transmembrane glycoprotein complexes. Each complex consists of four alpha subunits around the central channel and a beta l and beta 2 subunit peripherally dNA cloning has identified three types in central neurons(l, II and Ill), ml in skeletal muscle, and h and i in cardiac muscle Sodium channels are blocked by tetrodotoxin(produced by puffer fish),geo graphotoxin and by lipid-soluble amines used as local anaesthetic agents. They are activated by ciguatoxins, pyrethrin and low molecular weight polypeptide toxins from scorpions and sea anemones. e Chloride channels play an important role in stabilisation of the membrane potential, regulation of cell volume, transepithelial transport and secretion of fluid from secretory glands Activation of chloride channels can be achieved by Extracellular ligan Intracellular calcium Cyclic AMP proteins Mechanical stretch Transmembrane voltage Methods for the study of ion channel structure Physical conformation High resolution electron microscopy; Molecular structure Isolation of channel proteins by biochemical methods Molecular cloning to determine amino acid sequences of proteins Site-directed mutagenesis to alter sequences at selected sites Expression of channel proteins in host cells, e.g. Xenopus oocytes lon channel disorders Calcium channels: malignant hyperthermia Chloride channel: cystic fibrosis Sodium channels: Liddle's syndrome(aldosterone-activated sodium channels in the coll Mutant sodium channels: long QT syndrome; Brugada syndrome Potassium channels: isolated deafness syndrome
Leak channels, with an apparent lack of gating control. * Sodium channels are discrete, four domain, transmembrane glycoprotein complexes. Each complex consists of four alpha subunits around the central channel and a beta 1 and beta 2 subunit peripherally. DNA cloning has identified three types in central neurons (I, II and III), m1 in skeletal muscle, and h and I in cardiac muscle. Sodium channels are blocked by tetrodotoxin (produced by puffer fish), geographotoxin and by lipid-soluble amines used as local anaesthetic agents. They are activated by ciguatoxins, pyrethrin and low molecular weight polypeptide toxins from scorpions and sea anemones. * Chloride channels play an important role in stabilisation of the membrane potential, regulation of cell volume, transepithelial transport and secretion of fluid from secretory glands. Activation of chloride channels can be achieved by: Extracellular ligands Intracellular calcium Cyclic AMP G-proteins Mechanical stretch Cell swelling Transmembrane voltage Methods for the study of ion channel structure Physical conformation High resolution electron microscopy; Electron diffraction; Molecular structure Isolation of channel proteins by biochemical methods; Molecular cloning to determine amino acid sequences of proteins; Site-directed mutagenesis to alter sequences at selected sites; Expression of channel proteins in host cells, e.g. Xenopus oocytes. Ion channel disorders Calcium channels: malignant hyperthermia Chloride channel: cystic fibrosis Sodium channels: Liddle’s syndrome (aldosterone-activated sodium channels in the collecting ducts) Mutant sodium channels: long QT syndrome; Brugada syndrome Potassium channels: isolated deafness syndrome Cell membrane transport 11
■ Subcellular organelles Nucleus The nucleus is the most prominent cellular organelle, and contains the cells database or genome, which is encoded in DNA. It comprises the following CO Nuclear envelope Outer membrane, which is continuous with the endoplasmic reticulum Inner membrane Pore complex: nuclear pores connect the inner and outer membranes. This is a gated channel through which ribonucleoproteins are transported to the cytoplasm Nucleolus: which is associated with ribonucleic acid(RNA) processing and ribosome synthesis. It is not bounded by a membrane Fibrous matrix the structural skeleton of the nucleus Deoxyribonucleic acid (DNA)-protein complex: The nucleus contains all chromosomal DNA, the genetic material of the cell Functions of the nucleus The functions of the nucleus include Synthesis of new DNA, involving DNA replication during mitosis Synthesis of ribosomal RNA, messenger RNA, and transfer RNA. The nucleolus is the site of messenger RNA transcription and processing, and of ribosome a key role in cell division Endoplasmic reticulum The endoplasmic reticulum comprises two morphologically distinct systems that are interconnected to form a single membrane system. The functions of the system broadly consist of synthesis, storage, transport and detoxification. The endoplasmic reticulum consists of membrane-enclosed branching tubules and flattened sacs(cisternae)and vesicles, which intercommunicate throughou the cytosol forming an intracellular transport network. The cisternal space is enclosed within The endoplasmic reticulum is the largest subcellular organelle The rough endoplasmic reticulum contains ribosomes and is the site of protein synthesis, being involved in translation of messenger RNA into protein
& Subcellular organelles Nucleus The nucleus is the most prominent cellular organelle, and contains the cell’s database or genome, which is encoded in DNA. It comprises the following components: Nuclear envelope. Outer membrane, which is continuous with the endoplasmic reticulum membrane. Inner membrane. Pore complex: nuclear pores connect the inner and outer membranes. This is a gated channel through which ribonucleoproteins are transported to the cytoplasm. Nucleolus: which is associated with ribonucleic acid (RNA) processing and ribosome synthesis. It is not bounded by a membrane. Fibrous matrix: the structural skeleton of the nucleus. Deoxyribonucleic acid (DNA)-protein complex: The nucleus contains all chromosomal DNA, the genetic material of the cell. Functions of the nucleus The functions of the nucleus include: Synthesis of new DNA, involving DNA replication during mitosis. Synthesis of ribosomal RNA, messenger RNA, and transfer RNA. The nucleolus is the site of messenger RNA transcription and processing, and of ribosome assembly. A key role in cell division. Endoplasmic reticulum The endoplasmic reticulum comprises two morphologically distinct systems that are interconnected to form a single membrane system. The functions of the system broadly consist of synthesis, storage, transport and detoxification. The endoplasmic reticulum consists of membrane-enclosed branching tubules and flattened sacs (cisternae) and vesicles, which intercommunicate throughout the cytosol forming an intracellular transport network. The cisternal space is enclosed within. The endoplasmic reticulum is the largest subcellular organelle. The rough endoplasmic reticulum contains ribosomes and is the site of protein synthesis, being involved in translation of messenger RNA into protein. Cell physiology 12
Functions of smooth endoplasmic reticulum Site of mixed function oxidase systems in the liver, which detoxify various organic compounds The site of glucose release from glucose-6-phosphate in the liver via glucose 6-phosphatase. In muscle it forms the T-system of sarcoplasmic reticulum, which is involved in calcium sequestration. The smooth endoplasmic reticulum does not contain ribosomes and is the site of lipid (fatty acids and phospholipids) and steroid synthesis. This includes the synthesis of steroid hormones in the adrenal cortex and the gonads Golgi apparatus molecules. There are three levels of organisation. the modification of complex Cisternae: flattened sac-like membranes Dictyosomes: stacks of cis Golgi complex: an association of dictyosomes A Golgi apparatus usually contains a stack of four to six cisternae with an entry (cis)and an exit(trans)surface. The Golgi apparatus distributes newly synthesised proteins and lipids from the endoplasmic reticulum to the plasma mem- brane, lysosomes and secretory vesicles for export. It is concerned with po translational modification of proteins, including: O-glycosylation to form glycoprotein Proteoglycan formation; Sulphation of sugars and tyrosine residues. The effects of Golgi apparatus function involve the sorting of molecules for transport out of the cell by exocytosis, incorporation in the cell membrane as integral membrane proteins, or intracellular transport to lysosomes for digesti Mitochondria These are the respiratory organelles of the cell Mitochondria are about 2 um long and 0.5-1 um in diameter. A mitochondrion comprise e A smooth outer membrane, which recognises and translocates mitochondrial Ahighly folded inner membrane with invaginations, called cristae, projecting into the inner matrix. The spaces between the cristae are narrow. The inner
The smooth endoplasmic reticulum does not contain ribosomes and is the site of lipid (fatty acids and phospholipids) and steroid synthesis. This includes the synthesis of steroid hormones in the adrenal cortex and the gonads. Golgi apparatus The Golgi apparatus is an organelle involved in the modification of complex molecules. There are three levels of organisation: Cisternae: flattened sac-like membranes Dictyosomes: stacks of cisternae Golgi complex: an association of dictyosomes A Golgi apparatus usually contains a stack of four to six cisternae with an entry (cis) and an exit (trans) surface. The Golgi apparatus distributes newly synthesised proteins and lipids from the endoplasmic reticulum to the plasma membrane, lysosomes and secretory vesicles for export. It is concerned with posttranslational modification of proteins, including: O-glycosylation to form glycoproteins; Proteoglycan formation; Sulphation of sugars and tyrosine residues. The effects of Golgi apparatus function involve the sorting of molecules for transport out of the cell by exocytosis, incorporation in the cell membrane as integral membrane proteins, or intracellular transport to lysosomes for digestion. Mitochondria These are the respiratory organelles of the cell. Mitochondria are about 2 mm long and 0.5–1 mm in diameter. A mitochondrion comprises: * A smooth outer membrane, which recognises and translocates mitochondrial proteins. * A highly folded inner membrane with invaginations, called cristae, projecting into the inner matrix. The spaces between the cristae are narrow. The inner Functions of smooth endoplasmic reticulum Site of mixed function oxidase systems in the liver, which detoxify various organic compounds. The site of glucose release from glucose-6-phosphate in the liver via glucose 6-phosphatase. In muscle it forms the T-system of sarcoplasmic reticulum, which is involved in calcium sequestration. Subcellular organelles 13
membrane contains five complexes of integral membrane proteins: nicotin- amide adenine dinucleotide (NAD)H dehydrogenase, succinate dehydrogen- ase, cytochrome c reductase, cytochrome c oxidase and ATP synthase. These are involved in oxidative phosphorylation An internal gel-like matrix space, which contains Enzymes of the tricarboxylic acid cycle Enzymes involved in beta oxidation of fatty acids; Enzymes involved in urea formation Ribosomes An inter-membrane space The mitochondria synthesise aTP by the process of oxidative phosphorylation Mitochondrial content is highest in cells with a high energy expenditure The mitochondrial genome consists of 5-10 identical circular double-strande DNA molecules, which encode for two ribosomal rNa molecules: 22 transfer rNA molecules; and 13 polypeptides of complexes L, Ill, IV and V of the respiratory chain. Mitochondria replicate, transcribe and translate their DNA independently of nuclear DNA. Mitochondrial DNA is inherited maternally and does not recom bine. each mitochondrion acts as an individual unit because there is no evidence of transfer of mitochondrial proteins or DNA between mitochondria. Thus a mixture of normal and mutant mitochondrial dna can coexist within the same cell Mitochondrial diseases are disorders of energy metabolism, including defects of pyruvate metabolism, the Krebs cycle, respiratory chain, oxidative phospho ylation and fatty acid oxidation Over 90% of mitochondrial disorders are caused by mutations in nuclear genes. Mitochondrial DNA, however, mutates more than ten times as frequently as nuclear DNA As mitochondrial dnA does not possess any introns, a random mutation affects a coding DNA sequence. Furthermore, mitochondrial DNA lacks protective histones or other effective repair systems Lysosomes Lysosomes are vesicular structures limited by a single smooth membrane, com- prising the intracellular digestive system. Their functions include ntracellular digestion of proteins, carbohydrates, lipids and nucleic acids. Autophagy: the destruction of unwanted subcellular organelles. Autolysis: the digestion of cells after death The handling of the products of receptor-mediated endocytosis. The release of enzymes by exocytosis in the extracellular environment to gest external material
membrane contains five complexes of integral membrane proteins: nicotinamide adenine dinucleotide (NAD)H dehydrogenase, succinate dehydrogenase, cytochrome c reductase, cytochrome c oxidase and ATP synthase. These are involved in oxidative phosphorylation. * An internal gel-like matrix space, which contains: Enzymes of the tricarboxylic acid cycle; Enzymes involved in beta oxidation of fatty acids; Enzymes involved in urea formation; Gluconeogenesis enzymes; Ribosomes. * An inter-membrane space. The mitochondria synthesise ATP by the process of oxidative phosphorylation. Mitochondrial content is highest in cells with a high energy expenditure. The mitochondrial genome consists of 5–10 identical circular double-stranded DNA molecules, which encode for two ribosomal RNA molecules; 22 transfer RNA molecules; and 13 polypeptides of complexes I, III, IV and V of the respiratory chain. Mitochondria replicate, transcribe and translate their DNA independently of nuclear DNA. Mitochondrial DNA is inherited maternally and does not recombine. Each mitochondrion acts as an individual unit, because there is no evidence of transfer of mitochondrial proteins or DNA between mitochondria. Thus a mixture of normal and mutant mitochondrial DNA can coexist within the same cell. Mitochondrial diseases are disorders of energy metabolism, including defects of pyruvate metabolism, the Krebs cycle, respiratory chain, oxidative phosphorylation and fatty acid oxidation. Over 90% of mitochondrial disorders are caused by mutations in nuclear genes. Mitochondrial DNA, however, mutates more than ten times as frequently as nuclear DNA. As mitochondrial DNA does not possess any introns, a random mutation affects a coding DNA sequence. Furthermore, mitochondrial DNA lacks protective histones or other effective repair systems. Lysosomes Lysosomes are vesicular structures limited by a single smooth membrane, comprising the intracellular digestive system. Their functions include: Intracellular digestion of proteins, carbohydrates, lipids and nucleic acids. Autophagy: the destruction of unwanted subcellular organelles. Autolysis: the digestion of cells after death. The handling of the products of receptor-mediated endocytosis. The release of enzymes by exocytosis in the extracellular environment to digest external material. Cell physiology 14