Ionic Channels in Cells and Model Systems

This book is based on a series of lectures for a course on ionic channels held in Santiago, Chile, on November 17-20, 1984. It is intended as a tutorial guide on the properties, function, modulation, and reconstitution of ionic channels, and it should be accessible to graduate students taking their first steps in this field. In the presentation there has been a deliberate emphasis on the spe­ cific methodologies used toward the understanding of the workings and function of channels. Thus, in the first section, we learn to "read" single­ channel records: how to interpret them in the theoretical frame of kinetic models, which information can be extracted from gating currents in re­ lation to the closing and opening processes, and how ion transport through an open channel can be explained in terms of fluctuating energy barriers. The importance of assessing unequivocally the origin and purity of mem­ brane preparations and the use of membrane vesicles and optical tech­ niques in the stUGY of ionic channels are also discussed in this section. The patch-clamp technique has made it possible to study ion channels in a variety of different cells and tissues not amenable to more conven­ tional electrophysiological methods. The second section, therefore, deals with the use of this technique in the characterization of ionic channels in different types of cells, ranging from plant protoplasts to photoreceptors.

I. Methodologies.- 1 Kinetic Models and Channel Fluctuations.- 1. Introduction.- 2. Two-State Channel.- 3. Two Two-State Channels.- 4. Three-State Channel with Three Conductances.- 5. Three-State Channel with Only Two Conductances.- References.- 2 Single-Channel Currents and Postsynaptic Drug Actions.- 1. Introduction.- 2. Channel Gating as a Stochastic Process.- 3. Postsynaptic Channels in the Presence of Drugs.- 4. Reconstructing the Postsynaptic Current.- 5. Macroscopic and Molecular Consequences.- References.- 3 Voltage-Dependent Gating: Gating Current Measurement and Interpretation.- 1. Introduction.- 2. Voltage Gating.- 3. Gating Current Is a Capacitive Current.- 4. Measurement of Gating Currents.- 5. Gating of the Sodium Channel.- References.- 4 Characterizing the Electrical Behavior of an Open Channel via the Energy Profile for Ion Permeation: A Prototype Using a Fluctuating Barrier Model for the Acetylcholine Receptor Channel.- 1. Introduction.- 2. Theory.- 3. Confrontation with Experimental Data for the AChR Channel.- 4. Discussion.- References.- 5 The Use of Specific Ligands to Study Sodium Channels in Muscle.- 1. Introduction.- 2. Molecular Pharmacology of the Sodium Channel in Muscle.- 3. Sodium Channel in Cardiac Muscle: Are All Sodium Channels Alike?.- 4. Surface and Tubular Sodium Channels in Skeletal Muscle.- 5. Models for Sodium Channels in Muscle Membranes.- References.- 6 Isolation of Muscle Membranes Containing Functional Ionic Channels.- 1. Introduction.- 2. Excitation-Contraction Coupling.- 3. Ionic Channels and E-C Coupling.- 4. Isolation of Muscle Membranes.- 5. Concluding Remarks.- References.- 7 Methodologies to Study Channel-Mediated Ion Fluxes in Membrane Vesicles.- 1. Introduction.- 2. Channel-Mediated Tl+ Flux Measured by Fluorescence Quenching.- 3. Channel-Mediated Ion Fluxes Measured by Light Scattering.- References.- 8 Optical Studies on Ionic Channels in Intact Vertebrate Nerve Terminals.- 1. Introduction.- 2. Equivalence of Optical and Electrical Measurements of Membrane Potential.- 3. Optical Recording of Action Potentials from Nerve Terminals of the Frog Xenopus.- 4. Properties of the Action Potential in the Nerve Terminals.- 5. Ionic Basis of the Depolarizing Phase of the Action Potential.- 6. Concluding Remarks.- References.- 9 Optical Detection of ATP Release from Stimulated Endocrine Cells: A Universal Marker of Exocytotic Secretion of Hormones.- 1. Introduction.- 2. Methodological Considerations.- 3. Acetylcholine-Induced ATP Release from Chromaffin Cells: Calcium Dependence.- 4. Nicotinic Receptor Desensitization.- 5. Granular Nature of the Secreted ATP.- 6. ATP Release Evoked by Membrane Depolarization Is Mediated by Activation of Voltage-Gated Calcium Channels.- 7. ATP Release from Collagenase-Isolated Islets of Langerhans.- 8. Conclusion.- 9. Summary.- References.- II. Channels in Biological Membranes.- 10 Mechanotransducing Ion Channels.- 1. Introduction.- 2. Recording SA Channels.- 3. General Characteristics.- 4. Conductance Properties.- 5. Kinetic Properties.- 6. The Model.- 7. Comparing the Model to the Data.- 8. Future Prospects.- References.- 11 Ionic Channels in Plant Protoplasts.- 1. Introduction.- 2. Some Methodological Considerations.- 3. Voltage-Dependent Channels Opened by Hyperpolarization.- 4. Channels Affected by TEA.- 5. Conclusions.- References.- 12 Channels in Kidney Epithelial Cells.- 1. Introduction.- 2. Cell Culture.- 3. Patch-Clamp Methodology.- 4. Potassium Channel Characteristics.- 5. Channel Modulation.- 6. Conclusions.- References.- 13 Channels in Photoreceptors.- 1. Introduction.- 2. Vertebrate Photoreceptors.- 3. Invertebrate Photoreceptors.- References.- 14 Inactivation of Calcium Currents in Muscle Fibers from Balanus.- 1. Introduction.- 2. Methodological Considerations.- 3. Characteristics of Inward Currents.- 4. Mechanism of Inactivation.- References.- 15 Electrophysiological Studies in Endocrine Cells.- 1. Introduction.- 2. Whole-Cell Patch-Clamp Methodology.- 3. Cell Culture.- 4. Ionic Currents in GH3 Cells.- 5. Characteristics of Calcium Channels.- 6. Conclusions.- References.- III. Ionic Channel Reconstitution.- 16 Ion Channel Reconstitution: Why Bother?.- 1. Introduction and Background.- 2. Unexpected Surprises.- 3. Unconstrained Variables.- 4. Unrealized Hopes.- References.- 17 From Brain to Bilayer: Sodium Channels from Rat Neurons Incorporated into Planar Lipid Membranes.- 1. Perspectives and Background.- 2. Electrophysiology without Cells.- 3. A Closer Look at Batrachotoxin-Activated Sodium Channels in Bilayer Membranes.- 4. Looking Ahead.- References.- 18 Ionic Channels in the Plasma Membrane of Sea Urchin Sperm.- 1. Introduction.- 2. Are There Channels in Sea Urchin Sperm?.- 3. Reconstitution Studies with Isolated Sea Urchin Sperm Plasma Membrane.- 4. Channels in the Plasma Membrane of Sea Urchin Sperm: Implications for the Acrosome Reaction.- 5. Are There Receptors to the Egg Jelly in the Sea Urchin Sperm Plasma Membranes?.- 6. Perspectives.- References.- 19 Characterization of Large-Unitary-Conductance Calcium-Activated Potassium Channels in Planar Lipid Bilayers.- 1. Introduction.- 2. Channel Gating.- 3. Channel Conductance and Selectivity.- 4. Conductance of the Calcium-Activated K+ Channels.- 5. Selectivity of the Ca-K Channels.- 6. Blockade of the Ca-K Channels.- 7. Conclusions.- References.- IV. Ionic Channel Modulation.- 20 Metabolic Regulation of Ion Channels.- 1. Introduction.- 2. Second Messengers.- 3. Protein Phosphorylation.- 4. Summary.- References.- 21 The Cell-to-Cell Membrane Channel: Its Regulation by Cellular Phosphorylation.- 1. Introduction.- 2. The Cell-to-Cell Channels Are Up-Regulated by cAMP-Dependent Phosphorylation.- 3. The Cell-to-Cell Channels are Down-Regulated by Tyrosine Phosphorylation.- References.- 22 The ?-Cell Bursting Pattern and Intracellular Calcium.- 1. Introduction.- 2. Role of [Ca2+]i Dependence on Glucose.- 3. A Biophysical/Mathematical Model.- 4. Burst Frequency Depends on the Ratio [free Ca2+]i/[total Ca]i.- 5. Summary.- References.- 23 Neurotrophic Effects of in Vitro Innervation of Cultured Muscle Cells. Modulation of Ca2+-Activated K+ Conductances.- 1. Introduction.- 2. Methodological Considerations.- 3. Innervation and Muscle Cell Electrical Activity.- 4. Conclusions.- References.- V. Ionic Channel Structure, Functions, and Models.- 24 Correlation of the Molecular Structure with Functional Properties of the Acetylcholine Receptor Protein.- 1. Introduction.- 2. The AChR Macromolecule.- 3. Arrangement of Subunits in the AChR Macromolecule.- 4. The AChR Primary Structure, cDNA Recombinant Techniques, and Modeling Receptor Structure.- 5. Immunochemistry of AChR and the Testing of Models.- 6. Voltage-Gated and Agonist-Gated Channels: A Comparison.- 7. Dynamics of AChR and Lipids in the Membrane.- 8. Acetylcholine-Receptor-Controlled Channel Properties.- References.- 25 Amiloride-Sensitive Epithelial Sodium Channels.- 1. Introduction.- 2. Amiloride-Sensitive Na+ Transport Processes.- 3. Characterization of Amiloride-Sensitive Na+ Channels in Intact Epithelia.- 4. Incorporation of Amiloride-Sensitive Na+ Channels into Planar Bilayers.- 5. Concluding Remarks.- References.- 26 A Channel Model for Development of the Fertilization Membrane in Sea Urchin Eggs.- 1. Introduction.- 2. Processes Following Fertilization.- 3. Experimental Basis for Model.- 4. Description of Model.- 5. Equations of Model.- 6. Solutions of Model Equations.- References.