Premed · Premed · Cell Biology
Lecture 6: Membrane Transport: Ion Channels and Pumps
Cell Biology
Learning Objectives
By the end of this lecture, students will be able to:
- Classify ion channels by gating mechanism and ion selectivity
- Explain the molecular basis of ion selectivity in potassium channels
- Describe the generation and propagation of action potentials
- Explain the mechanism of synaptic transmission at chemical synapses
- Discuss the pharmacology of ion channels and their clinical relevance
Lecture Content
I. Ion Channel Properties and Classification
Ion channels are transmembrane proteins that form aqueous pores through which specific ions can flow rapidly across the membrane. They are characterized by three key properties. Ion selectivity refers to the preferential passage of specific ions such as Na+, K+, Ca2+, or Cl-. Gating describes the ability of channels to open and close in response to specific stimuli. Conductance measures the rate of ion flow when the channel is open and is expressed in picosiemens (pS), with typical single-channel conductances ranging from about 1 to 300 pS.
Ion channels can be classified by their gating mechanism. Voltage-gated channels open and close in response to changes in membrane potential. Voltage-gated Na+ channels (Nav) drive the depolarization phase of action potentials, voltage-gated K+ channels (Kv) mediate repolarization, and voltage-gated Ca2+ channels (Cav) trigger neurotransmitter release and muscle contraction. Ligand-gated channels (ionotropic receptors) open upon binding of a specific extracellular ligand. The nicotinic acetylcholine receptor (nAChR) is a Na+/K+ channel, the GABA-A receptor is an inhibitory Cl- channel, and glutamate receptors (AMPA, NMDA, and kainate types) are excitatory Na+/Ca2+ channels. Mechanically-gated channels respond to mechanical forces such as stretch or pressure; Piezo channels mediate touch sensation in mammals, while MscL channels in bacteria respond to osmotic stress. Leak channels, such as the K+ leak channels TREK and TASK, are constitutively open and play a critical role in maintaining the resting membrane potential.
Channels can also be classified by ion selectivity into cation channels (selective for Na+, K+, or Ca2+, or non-selective) and anion channels (Cl- channels of the ClC family and CFTR).
II. Structure and Selectivity of Ion Channels
The structural basis of ion selectivity was revealed when Roderick MacKinnon (Nobel Prize, 2003) solved the crystal structure of the KcsA potassium channel from Streptomyces lividans. KcsA is a tetramer of identical subunits, each containing two transmembrane helices (TM1 and TM2) connected by a pore loop (P-loop). The selectivity filter is formed by the signature sequence GYG (Gly-Tyr-Gly) in the P-loop, where backbone carbonyl oxygens line the narrowest part of the pore. K+ ions are precisely coordinated by these carbonyls, which substitute for the water molecules of K+'s hydration shell. Na+, despite being smaller than K+, cannot be properly coordinated in this configuration because shedding its hydration shell within the filter is energetically unfavorable. Two K+ ions occupy the filter simultaneously, and the electrostatic repulsion between them drives rapid throughput. The channel gate is formed by the crossing of TM2 helices at the cytoplasmic face of the channel. The transport rate approaches the diffusion limit at approximately 10^8 ions per second.
Voltage-gated channels have a more complex architecture, with each subunit containing six transmembrane segments (S1-S6). The S1-S4 segments form the voltage-sensing domain, with S4 containing positively charged arginine or lysine residues at every third position. Upon depolarization, the S4 segment moves outward, opening the pore formed by S5, S6, and the intervening P-loop. Nav channels are single polypeptides comprising four homologous domains (I-IV), each with six transmembrane segments. They undergo fast inactivation through a "ball-and-chain" mechanism: the intracellular loop between domains III and IV contains an Ile-Phe-Met motif that plugs the pore within milliseconds of opening.
<image>Structure of the KcsA potassium channel and its selectivity filter. Panel A: Top-down view showing the tetrameric arrangement of four subunits around a central pore. Panel B: Side view cross-section showing two subunits with TM1 and TM2 helices, the P-loop forming the selectivity filter, and the bundle crossing gate at the bottom. Panel C: Close-up of the selectivity filter showing two K+ ions coordinated by backbone carbonyl oxygens of the GYG motif, with a diagram showing why Na+ (too small) cannot be properly coordinated and is rejected. Water molecules alternate with K+ ions in the filter.</image>
III. The Action Potential
The action potential is the rapid, transient reversal of membrane potential that serves as the electrical signal in excitable cells. In the resting state, the membrane potential sits near -70 mV, maintained by K+ leak channels and the Na+/K+-ATPase.
During the depolarization phase, an initial stimulus opens some Na+ channels, allowing Na+ influx that depolarizes the membrane. If this depolarization reaches the threshold (approximately -55 mV), voltage-gated Nav channels open in a self-reinforcing process: depolarization opens more Nav channels, which causes more Na+ influx, which causes more depolarization. This positive feedback drives the membrane potential rapidly toward E_Na (approximately +60 mV), with the membrane briefly overshooting to approximately +30 to +40 mV.
Repolarization begins when Nav channels inactivate within about 1 millisecond through fast inactivation, and voltage-gated Kv channels open with a slight delay. The resulting K+ efflux drives the membrane potential back toward E_K. A brief hyperpolarization (undershoot) occurs because Kv channels remain open slightly longer than needed, pushing the membrane potential below the resting value to approximately -80 mV before they close and the membrane returns to rest.
The action potential exhibits two refractory periods. During the absolute refractory period, Nav channels are inactivated and no action potential can be generated regardless of stimulus strength. During the relative refractory period, some Nav channels have recovered, but a larger-than-normal stimulus is required.
Action potentials have distinctive properties: they are all-or-none responses, they propagate without decrement along the axon through local current spread, and in myelinated neurons they exhibit saltatory conduction, jumping between nodes of Ranvier. Propagation speeds range from 1 to 100 m/s depending on axon diameter and the degree of myelination.
<image>The action potential and underlying ion channel activity. Panel A: Voltage trace of an action potential showing resting potential (-70 mV), threshold (-55 mV), depolarization phase rising to +40 mV, repolarization, hyperpolarization undershoot, and return to resting potential. Time scale ~2-3 ms. Panel B: Below the voltage trace, two conductance curves — Na+ conductance (rapid rise and fall during depolarization) and K+ conductance (delayed rise during repolarization). Panel C: Diagrams of Nav channel states: closed (resting), open (activated), inactivated (ball-and-chain blocking pore), and recovered (ready to open again).</image>
IV. Synaptic Transmission
At chemical synapses, an electrical signal is converted into a chemical signal and then back into an electrical signal. The presynaptic terminal is separated from the postsynaptic cell by a synaptic cleft approximately 20 nm wide. When an action potential arrives at the presynaptic terminal, voltage-gated Ca2+ channels open and Ca2+ floods into the terminal. This Ca2+ influx triggers the fusion of neurotransmitter-filled synaptic vesicles with the presynaptic membrane through the action of SNARE proteins and the Ca2+ sensor synaptotagmin. Neurotransmitter is released into the cleft by exocytosis and binds to ligand-gated ion channels on the postsynaptic membrane.
At excitatory synapses, neurotransmitter binding produces a depolarizing excitatory postsynaptic potential (EPSP). Glutamate activates AMPA receptors (causing Na+ influx) and NMDA receptors (allowing Na+ and Ca2+ influx). At inhibitory synapses, the result is a hyperpolarizing inhibitory postsynaptic potential (IPSP), generated when GABA activates GABA-A receptors (Cl- influx) or glycine activates glycine receptors (also Cl- influx). Neurotransmitter signaling is terminated by enzymatic degradation (as with acetylcholinesterase), reuptake transporters, or simple diffusion away from the cleft.
The neuromuscular junction is a specialized synapse between a motor neuron and a skeletal muscle fiber. Acetylcholine released by the motor neuron binds nicotinic ACh receptors on the muscle end plate, producing Na+ influx, an end-plate potential, a muscle action potential, and ultimately muscle contraction. Curare blocks this process by competitively antagonizing the nAChR, causing paralysis. Botulinum toxin prevents ACh release entirely by cleaving SNARE proteins.
V. Patch Clamp Electrophysiology
The patch clamp technique, developed by Erwin Neher and Bert Sakmann (Nobel Prize, 1991), revolutionized the study of ion channels by enabling the recording of currents through individual channel molecules. A glass micropipette is pressed against the cell membrane to form an extremely tight seal (gigaohm seal), which electrically isolates a small patch of membrane.
Four configurations are possible. The cell-attached configuration records single channels in an intact cell. The whole-cell configuration, formed by rupturing the patch, records the total current across the entire cell membrane. The inside-out configuration creates an excised patch with the cytoplasmic face exposed to the bath solution, while the outside-out configuration exposes the extracellular face. This technique reveals single-channel conductance, open probability, kinetics, and pharmacological properties, and can resolve currents as small as approximately 1 picoampere.
VI. Channelopathies and Pharmacology
Channelopathies are diseases caused by mutations in ion channel genes. Cystic fibrosis results from mutations in the CFTR Cl- channel, with the delta-F508 mutation (causing impaired protein folding) being the most common. Long QT syndrome arises from mutations in cardiac K+ or Na+ channels and predisposes to dangerous arrhythmias. Epilepsy can result from mutations in Nav, Kv, or GABA receptor channels. Myotonia involves mutations in skeletal muscle Cl- or Na+ channels. Liddle syndrome is a gain-of-function mutation in the epithelial sodium channel (ENaC) that causes hypertension.
Ion channels are major pharmacological targets. Local anesthetics such as lidocaine block Nav channels. Tetrodotoxin from pufferfish blocks Nav channels extracellularly. Benzodiazepines like diazepam enhance GABA-A receptor Cl- conductance. Dihydropyridines such as nifedipine block L-type Cav channels and serve as antihypertensives. Sulfonylureas like glibenclamide block K-ATP channels in pancreatic beta cells, stimulating insulin release for diabetes management.
