Lecture Notes on Excitable Tissues PDF

LECTURE NOTE 1 FOR 200L By MRS NZOPUTAM O.J. Excitable and contractile 2 tisssues, resting membrane potential and action potential  Excitable Tissues 3 u Tissues which are capable of generation and transmission of elec...

LECTURE NOTE 1 FOR 200L By MRS NZOPUTAM O.J. Excitable and contractile 2 tisssues, resting membrane potential and action potential  Excitable Tissues 3 u Tissues which are capable of generation and transmission of electrochemical impulses along the membrane are known as excitable tissues. u These include 1. Nerve 2. cardiac muscle 3. skeletal muscle and 4. smooth muscle uNon-excitable are 4 RBC Intestinal cells Fibroblasts Adipocytes  Resting Membrane Potential 5 u The resting membrane potential is the potential difference that exists across the membrane of excitable cells at rest (i.e. in the period between action potentials) u The resting membrane potential is established by diffusion potentials, which result from the concentration differences for various ions across the cell membrane. u The resting membrane potential of excitable cells falls in the range of -70 to -90 mV. uThese values can best be explained by the 6 concept of relative permeabilities of the cell membrane. uThe resting membrane potential is close to the equilibrium potentials for K+ and Cl- because the permeability to these ions at rest is high. uWhile it is far from the equilibrium potentials for Na+ and Ca2+ because the permeability to these ions at rest is low. uExcitable tissues have more 7 negative RMP ( - 70 mV to - 90 mV) u Non-excitable tissues have less negative RMP  -53 mV epithelial cells  -8.4 mV RBC  -20 to -30 mV fibroblasts  -58 mV adipocytes  Resting Membrane Potential 8 depends on following factors u – Ionic distribution across the membrane – Membrane permeability – Other factors Na+/K+ pump Ionic distribution 9 Major ions ion intrcellular extracellular u– Extracellular ions Na+ 10 142 Sodium, Chloride K+ 140 4 u– Intracellular ions Potassium, Proteinate Cl- 4 103 Ca2+ 0 2.4 HC03- 10 28  membrane permeability 10 Gibbs Donnan Equilibrium uW h e n t wo s o l u t i o n s c o n t a i n i n g i o n s a r e separated by membrane that is permeable to some of the ions and not to others an electrochemical equilibrium is established uElectrical and chemical energies on either side of the membrane are equal and opposite to each other  Flow of Potassium 11 u Intracellular Potassium concentration is more than extracellular u Membrane is freely permeable to K+ u There is therefore an efflux of K+ u Entryof positive ions in to the extracellular fluid creates positivity outside and negativity inside u Outside positivity then resists efflux of K+ (since K+ is a positive ion) At a certain voltage an equilibrium is reached and K+ efflux stops u This is known as Nernst potential or equilibrium potential  Nernst potential (Equilibrium potential) 12 u is the potential level across the membrane that will exactly prevent net diffusion of an ion Nernst equation determines this potential. ion intrcellular extracellular Nernst potential Na+ 10 142 +58 K+ 140 4 -92 Cl- 4 103 -89 Ca2+ 0 2.4 +129 HC03- 10 28 -23 (mmol/l)  Goldman Equation u When the membrane is permeable to several ions 13 the equilibrium potential that develops depends on – Polarity of each ion – Membrane permeability – Ionic conc In the resting state – K+ permeability is 20 times more than that of Na+ Leaky channels (K-Na leak channel) – More permeable to K  Na/K pump 14  Is an Active transport system for Na+-K+ exchange using energy u the Na+-K+ ATPase play important roles, in creating the resting membrane potential u First, there is a small direct electrogenic contribution of the Na+-K+ ATPase, since 3 Na+ efflux is coupled with 2 K+ influx i.e. three Na+ ions is pumped out of the cell for every two K+ ions pumped into the cell. u Second,the more important indirect contribution is in maintaining the concentration gradient for K+ across the15 cell membrane, u which then is responsible for the K+ diffusion potential that drives the membrane potential toward the K+ equilibrium potential. u With the net effect of causing negative charge inside the membrane u Thus, the Na+-K+ ATPase is necessary to create and maintain the K+ concentration gradient, which establishes the resting membrane potential. (A similar argument can be made for the role of the Na+-K+ ATPase in the upstroke of the action potential, where it maintains the ionic gradient for Na+ across the cell  Factors contributing to RMP  One of the main factors is K+ efflux (Nernst 16 Potential: - 92mV)  Contribution of Na influx is little (Nernst Potential: +61mV)  Na/K pump which causes more negativity inside the membrane  Negatively charged protein ions remaining inside the membrane contributes to the negativity u Net result: -70 to -90 mV inside  Electrochemical gradient 17 u At this electrochemical equilibrium, there is an exact balance between two opposing forces: 1. Chemical driving force = ratio of concentrations on 2 sides of membrane (concentration gradient) The concentration gradient that causes K+ to move from inside to outside taking along positive charge and 2. Electrical driving force = potential difference across membrane Opposing electrical gradient that increasingly tends to stop K+ from moving across the membrane 3. Equilibrium: when chemical driving force is balanced by electrical driving force  18 ACTION POTENTIAL uAction potential is a phenomenon of excitable 19 cells, such as nerve and muscle, uit consists of a rapid depolarization (upstroke) followed by repolarization of the membrane potential. u Action potentials are the basic mechanism for transmission of information in the nervous system and in all types of muscle.  Terminology used for discussion of the action potential, the refractory periods, 20 and the propagation of action potentials: u Depolarization is the process of making the membrane potential less negative. As noted, the usual resting membrane potential of excitable cells is oriented with the cell interior negative. Depolarization makes the interior of the cell less negative, or it may even cause the cell interior to become positive u Hyperpolarizationis the process of making the membrane potential more negative.  Terminology cont…….s 21 u Inward current is the flow of positive charge into the cell. Thus, inward currents depolarize the membrane potential. An example of an inward current is the flow of Na+ into the cell during the upstroke of the action potential. u Outward current is the flow of positive charge out of the cell. Outward currents hyperpolarize the membrane potential. An example of an outward current is the flow of K+ out of the cell during the repolarization phase of the action potential. Terminology cont…….s uThreshold potential is the membrane potential 22 at which occurrence of the action potential is inevitable.  Because the threshold potential is less negative than the resting membrane potential, an inward current is required to depolarize the membrane potential to threshold  If net inward current is less than net outward current, the membrane will not be depolarized to threshold, and no action potential will occur (see all-or-none response).  Terminology cont…….s 23 u Overshoot is that portion of the action potential where the membrane potential is positive (cell interior positive). u Undershoot, or hyperpolarizing after potential, is that portion of the action potential, following repolarization, where the membrane potential is actually more negative than it is at rest. u Refractoryperiod is a period during which another normal action potential cannot be elicited in an excitable cell. Refractory periods can be absolute or relative.  CHARACTERISTICS OF ACTION POTENTIALS 24 uAction potentials have three basic characteristics: stereotypical size and shape, propagation, and all- or-none response. 1. Stereotypical size and shape: Each normal action potential for a given cell type looks identical, depolarizes to the same potential, and repolarizes back to the same resting potential. 2. Propagation: An action potential at one site causes depolarization at adjacent sites, bringing those adjacent sites to threshold. 3. All-or-none response: An action potential either occurs or does not occur. 25 a. If an excitable cell is depolarized to threshold in a normal manner, then the occurrence of an action potential is inevitable. ie Once the threshold level is reached, AP is set off and no one can stop it - Like a gun b. On the other hand, if the membrane is not depolarized to threshold, no action potential can occur. Ie Until the threshold level the potential is graded c. indeed, if the stimulus is applied during the refractory period, then either no action potential occurs, or the action potential will occur but not have the stereotypical size and shape.  Principle of All or none law u The principle of All or none law says that the strength by which a nerve or muscle fiber responds to a stimulus is not dependent on the strength of the stimulus u If the stimulus is any strength above threshold, the nerve or muscle fiber will give a complete response or otherwise no response at all  Physiological basis of Action Potential in the nerve. 27 Inside of the membrane is Negative During RMP and Positive When an AP is generated 1. Initially membrane is slowly depolarised Until the threshold level is reached (This may be caused by the stimulus) When the threshold level is reached – Voltage-gated Na+ channels open up – Since Na conc outside is more than the inside – Na influx will occur – Positive ion coming inside increases the positivity of the membrane potential and causes depolarisation 2. This depolarization, When it reaches +30, Na+ channels close – Then Voltage-gated K+ channels open up 28 – K+ efflux occurs – Positive ion leaving the inside causes more negativity inside the membrane – Repolarisation occurs 3. When reaching the Resting level, rate slows down 4. Can go beyond the resting level this is known as hyperpolarisation 5. Since Na+ has come in and K+ has gone out 29  Membrane has become negative  But ionic distribution has become unequal 6. Na+/K+ pump restores Na+ and K+ conc slowly – By pumping 3 Na+ ions outward and 2+ K ions inward u Na+ channel – This has two gates 30 Activation and inactivation gates At rest: the activation gate is closed At threshold level: activation gate opens – Na+ influx will occur – Na+ permeability increases to 500 fold when reaching +30, inactivation gate closes – Na influx stops Inactivation gate will not reopen until resting membrane potential is reached Na+ channel opens fast uK+ channel 31 – This has only one gate At rest: K+ channel is closed – At +30, K+ channel open up slowly This slow activation causes K efflux – After reaching the resting still slow K+ channels may remain open: causing further hyperpolarisation  Note 32 uSpike potential means Sharp upstroke and downstroke u Time duration of AP in the nerve is 1 msec Summary of nerve action potential 33  REFRACTORY PERIODS 34 uDuring the refractory periods, excitable cells are incapable of producing normal action potentials. uThe refractory period includes 1. an absolute refractory period and 2. a relative refractory period  Absolute Refractory Period: 35 u This overlaps with almost the entire duration of the action potential. u Duringthis period, no matter how great the stimulus, another action potential cannot be elicited. u Thebasis for the absolute refractory period is closure of the inactivation gates of the Na+ channel in response to depolarization. u These inactivation gates are in the closed position until the cell is repolarized back to the resting membrane potential u Relative Refractory Period u The relative refractory period begins at the end of 36 the absolute refractory period u and overlaps primarily with the period of the hyperpolarizing afterpotential. u During this period, an action potential can be elicited, but only if a greater than usual depolarizing (inward) current is applied. u The basis for the relative refractory period is the higher K+ conductance than is present at rest. u Because the membrane potential is closer to the K+ equilibrium potential, more inward current is needed to  PROPAGATION OF ACTION POTENTIALS 37  Propagation of action potentials down a ner ve or muscle fiber occurs by the spread of local currents from active regions to adjacent inactive regions. u At r est, the entir e ner ve axon is at the r esting membrane potential, with the cell interior negative. u Action potentials are initiated in the initial segment of the axon, nearest the nerve cell body. u They propagate down the axon by spread of local currents as shown in the diagram below. u PROPAGATION OF ACTION POTENTIALS 38  Conduction Velocity 39 u The speed at which action potentials are conducted along a nerve or muscle fiber is known as the conduction velocity. u This property is of great physiologic importance because it determines the speed at which information can be transmitted in the nervous system. u To understand conduction velocity in excitable tissues, two major concepts must be explained: the time constant and the length constant. These concepts, called cable properties, explain how nerves and muscles act as cables to conduct electrical activity. u The time constant indicates how quickly a cell membrane depolarizes in response to an inward 40 current or how quickly it hyperpolarizes in response to an outward current. Factors that affect time constant are 1. membrane resistance 2. membrane capacitance u The length constant indicates how far a depolarizing current will spread along a nerve. In other words, the longer the length constant, the farther the current spreads down the nerve fiber u Changes in Conduction Velocity u Two mechanisms increase conduction velocity along a nerve: 41 1. increasing the size of the nerve fiber and 2. myelinating the nerve fiber. u Increasing nerve diameter. Increasing the size of a nerve fiber increases conduction velocity ie the larger the fiber, the lower the internal resistance., but anatomic constraints limit how large nerves can become. Therefore, a second mechanism, myelination, is invoked to increase conduction velocity. u conduction of action potentials is faster in myelinated nerves than in unmyelinated nerves because action potentials "jump" long distances from one node to the next, a process called saltatory conduction. u Membrane stabilisers Membrane stabilisers (these decrease excitability) 42 Increased serum Ca++ – Hypocalcaemia causes membrane instability and spontaneous activation of nerve membrane – Reduced Ca level facilitates Na entry – Spontaneous activation Decreased serum K+ Local anaesthetics Acidosis Hypoxia u Membrane destabilisers (these increase excitability) Decreased serum Ca++ Increased serum K+ Alkalosis  Muscle action potentials 43 Skeletal muscle Smooth muscle Cardiac muscle u Skeletalmuscle Similar to nerve action potential 44 u Cardiac muscle action potential u This is made up of five Phases  phase 0: depolarization  phase 1: short repolarization  phase 2: plateau phase  phase 3: repolarization  phase 4: resting Duration is about 250 msec due to the plateau u Cardiac muscle action potential Phase 0: depolarisation 45 (Na+ influx through fast Na+ channels) Phase 1: short repolarisation (K+ efflux through K+ channels, Cl- influx as well) Phase 2: plateau phase (Ca ++ influx through slow Ca ++ channels) Phase 3: repolarisation (K + efflux through K + channels) Phase 4: resting u Smooth muscle 46 uResting membrane potential may be about - 55mV uAction potential is similar to nerve AP uBut AP is not necessary for its contraction uSmooth muscle contraction can occur by hormones Synaptic and Neuromuscular Transmission 47 uA synapse is a site where information is transmitted from one cell to another. The information can be transmitted either electrically (electrical synapse) or via a chemical transmitter (chemical synapse). TYPES OF SYNAPSES 1. Electrical Synapses 2. Chemical Synapses 1. Electrical synapses allow current to flow from one excitable cell to the next via low resistance pathways 48 between the cells called gap junctions. Gap junctions are found in cardiac muscle and in some types of smooth muscle and account for the very fast conduction in these tissues. For example, cardiac ventricular muscle, in the uterus, and in the bladder. 2. In chemical synapses, there is a gap between the presynaptic cell membrane and the postsynaptic cell membrane, known as the synaptic cleft. Information is transmitted across the synaptic cleft via a neurotransmitter, a substance that is released from the presynaptic terminal and binds to receptors on the postsynaptic terminal  SEQUENCE OF EVENTS THAT OCCUR AT 49 CHEMICAL SYNAPSES: 1. An action potential in the presynaptic cell causes Ca2 + channels to open. 2. An influx of Ca2+ into the presynaptic terminal causes the neurotransmitter, which is stored in synaptic vesicles, to be released by exocytosis. 3. The neurotransmitter diffuses across the synaptic cleft, binds to receptors on the postsynaptic membrane, and produces a change in membrane potential on the postsynaptic cell. u The change in membrane potential on the postsynaptic cell membrane can be either excitatory or inhibitory, depending 50 on the nature of the neurotransmitter released from the presynaptic nerve terminal. a. If the neurotransmitter is excitatory, it causes depolarization of the postsynaptic cell; b. if the neurotransmitter is inhibitory, it causes hyperpolarization of the postsynaptic cell. u In contrast to electrical synapses, neurotransmission across chemical synapses is unidirectional (from presynaptic cell to postsynaptic cell). The synaptic delay is the time required for the multiple steps in chemical neurotransmission to occur. NEUROMUSCULAR JUNCTION-EXAMPLE OF A CHEMICAL SYNAPSE 51 u Motor Units u Motoneurons are the nerves that innervate muscle fibers. u A motor unit comprises a single motoneuron and the muscle fibers it innervates. u Motor units vary considerably in size: a) A single motoneuron may activate a few muscle fibers or thousands of muscle fibers. Predictably, small motor units are involved in fine motor activities (e.g., facial expressions), and large motor units are involved in gross muscular activities (e.g., quadriceps muscles used in running). u Sequence of Events at the Neuromuscular Junction 52  Sequence of events in neuromuscular transmission 1. 53 Action potential travels down the motoneuron to the presynaptic terminal. 2. Depolarization of the presynaptic terminal opens Ca2+ channels, and Ca2+ flows into the terminal. 3. Acetylcholine (ACh) is extruded into the synapse by exocytosis. 4. ACh binds to its receptor on the motor end plate. 5. Channels for Na+ and K+ are opened in the motor end plate. 6. Depolarization of the motor end plate causes action potentials to be generated in the adjacent muscle tissue. 7. ACh is degraded to choline and acetate by acetylcholinesterase (AChE); choline is taken back into the presynaptic terminal on an Na + -choline cotransporter. uThe synapse between a 54 motoneuron and a muscle fiber is called the neuromuscular junction Synthesis and degradation of acetylcholine 55 u Agents Affecting Neuromuscular Transmission 56 u Botulinus toxin Blocks ACh release from presynaptic terminals causing total blockade, paralysis of respiratory muscles, and death u Curare Competes with ACh for receptors on motor end plate decreasing size of EPP; in maximal doses produces paralysis of respiratory muscles and death u Neostigmine AChE inhibitor (anticholinesterase) which prolongs and enhances action of ACh at motor end plate u Hemicholinium Blocks reuptake of choline into presynaptic terminal depleting ACh stores from presynaptic terminal u  TYPES OF SYNAPTIC ARRANGEMENTS u There 57 are several types of relationships between the presynaptic element and the postsynaptic element): 1. one-to-one- whereA single action potential in the presynaptic cell, the motoneuron, causes a single action potential in the postsynaptic cell, the muscle fiber. 2. one-to-many- uncommon, but found, at the synapses of motoneurons on Renshaw cells of the spinal cord. An action potential in the presynaptic cell, the motoneuron, causes a burst of action potentials in the postsynaptic cells. This arrangement causes amplification of activity. 3. many-to-one-very common, here an action potential in the presynaptic cell is insufficient to produce an action potential in the postsynaptic cell. Instead, many presynaptic cells converge on the postsynaptic cell, summate, to determines whether the u EXCITATORY AND INHIBITORY POSTSYNAPTIC POTENTIALS u 58 The many-to-one synaptic arrangement is a common configuration in which many presynaptic cells converge on a single postsynaptic cell, with the inputs being either excitatory or inhibitory u Excitatory postsynaptic potentials (EPSPs) are synaptic inputs that depolarize the postsynaptic cell, bringing the membrane potential closer to threshold and closer to firing an action potential.  They are produced by opening Na+ and K+ channels, similar to the nicotinic ACh receptor.  The membrane potential is driven to a value approximately halfway between the equilibrium potentials for Na+ and K+, or 0 mV, which is a depolarized state.  Excitatory neurotransmitters include ACh, norepinephrine, epinephrine, dopamine, glutamate, and serotonin. u Inhibitory postsynaptic potentials (IPSPs) are 59 synaptic inputs that hyperpolarize the postsynaptic cell, taking the membrane potential away from threshold and farther from firing an action potential.  IPSPs are produced by opening Cl- channels.  The membrane potential is driven toward the Cl- equilibrium potential (approximately -90 mV), which is a hyperpolarized state.  Inhibitory neurotransmitters are γ-aminobutyric acid (GABA) and glycine u INTEGRATION OF SYNAPTIC INFORMATION u The presynaptic information that arrives at the synapse may be integrated 60 in one of two ways, spatially or temporally. u Spatial summation occurs when two or more presynaptic inputs arrive at a postsynaptic cell simultaneously. u and if excitatory, will combine to produce greater depolarization than either input would produce separately. u But if one input is excitatory and the other is inhibitory, they will cancel each other out. u Spatial summation may occur, even if the inputs are far apart on the nerve cell body, because EPSPs and IPSPs are conducted so rapidly over the cell membrane. u Temporal summation occurs when two presynaptic inputs arrive at the postsynaptic cell in rapid succession. Because the inputs overlap in time, they summate. u Other Phenomena That Alter Synaptic Activity u Facilitation, augmentation, and posttetanic potentiation are phenomena 61that may occur at synapses. u In each instance, repeated stimulation causes the response of the postsynaptic cell to be greater than expected. u The common underlying mechanism is believed to be an increased release of neurotransmitter into the synapse, possibly caused by accumulation of Ca2+ in the presynaptic terminal. u Long-term potentiation occurs in storage of memories and involves both increased release of neurotransmitter from presynpatic terminals and increased sensitivity of postsynaptic membranes to the transmitter. u Synaptic fatigue may occur where repeated stimulation produces a smaller than expected response in the postsynaptic cell, possibly resulting from the depletion of neurotransmitter stores from the presynaptic terminal. u NEUROTRANSMITTERS u The transmission of information at chemical synapses involves 62the release of a neurotransmitter from a presynaptic cell, diffusion across the synaptic cleft, and binding of the neurotransmitter to specific receptors on the postsynaptic membrane to produce a change in membrane potential. u The following criteria are used to formally designate a substance as a neurotransmitter: 1. The substance must be synthesized in the presynaptic cell; 2. the substance must be released by the presynaptic cell upon stimulation; and 3. if the substance is applied exogenously to the postsynaptic membrane at physiologic concentration, the response of the postsynaptic cell must mimic the in vivo response. uNeurotransmitter substances can be grouped in 63 the following categories: 1. acetylcholine, 2. biogenic amines, 3. amino acids, and 4. neuropeptides  1. Acetylcholine u The role of acetylcholine (ACh) is vitally important. 64 1. ACh is the only neurotransmitter that is utilized at the neuromuscular junction. 2. It is the neurotransmitter released from all preganglionic and most postganglionic neurons in the parasympathetic nervous system and from all preganglionic neurons in the sympathetic nervous system. 3. It is also the neurotransmitter that is released from presynaptic neurons of the adrenal medulla. 2. Biogenic Amines: 65 Dopamine, Epinephrine, Histamine, Norepinephrine, Serotonin 3. Amino Acids: γ-Aminobutyric acid (GABA), Glutamate, Glycine 4. Neuropeptides: Adrenocorticotropin (ACTH), Cholecystokinin, Dynorphin , Endorphins, Enkephalins ,Glucose- dependent insulinotropic peptide (GIP), Glucagon , Neurotensin , Oxytocin, Secretin, Substance P , Thyrotropin-releasing hormone (TRH), Vasopressin, Vasoactive intestinal peptide (VIP)  Skeletal Muscle 66 u Contraction of skeletal muscle is under voluntary control. u Each skeletal muscle cell is innervated by a branch of a motoneuron. u Action potentials are propagated along the motoneurons, leading to release of ACh at the neuromuscular junction, depolarization of the motor end plate, and initiation of action potentials in the muscle fiber.  Excitation-contraction Coupling. u These are Events that occurs between the action potential 67 in the muscle fiber and contraction of the muscle fiber. u MUSCLE FILAMENTS u Each muscle fiber behaves as a single unit, is multinucleate, and contains myofibrils. u The myofibrils are surrounded by sarcoplasmic reticulum and are invaginated by transverse tubules (T tubules). u Each myofibril contains interdigitating thick and thin filaments, which are arranged longitudinally and cross-sectionally in sarcomeres u The repeating units of sarcomeres account for the unique banding pattern seen in striated muscle (which includes both skeletal and cardiac muscle). u The Thick filaments: u comprise a large molecular weight protein called myosin, which68 has six polypeptide chains, including one pair of heavy chains and two pairs of light chains u Most of the heavy-chain myosin has an α-helical structure, in which the two chains coil around each other to form the "tail" of the myosin molecule. u The four light chains and the N terminus of each heavy chain form two globular "heads" on the myosin molecule. u These globular heads have an actin-binding site, which is necessary for cross-bridge formation, and a site that binds and hydrolyzes ATP (myosin ATPase). u THE THIN FILAMENTS: u are composed of three proteins: actin, tropomyosin, and troponin Structure of thick (A) and thin (B) filaments of skeletal muscle 69 Troponin is a complex of three proteins: I, troponin I; T, troponin T; and C, troponin C u Actin is a globular protein and, in this globular form, is called G-actin. u In the thin filaments, G-actin is polymerized into two strands that70are twisted into an α-helical structure to form filamentous actin, called F-actin. u Actin has myosin-binding sites. u When the muscle is at rest, the myosin-binding sites are covered by tropomyosin so that actin and myosin cannot interact. u Tropomyosin is a filamentous protein that runs along the groove of each twisted actin filament. u At rest, its function is to block the myosin-binding sites on actin. u If contraction is to occur, tropomyosin must be moved out of the way so that actin and myosin can interact. u Troponin is a complex of three globular proteins (troponin T, troponin I, and troponin C) located at regular intervals along the 71 tropomyosin filaments. u Troponin T (T for tropomyosin) attaches the troponin complex to tropomyosin. u Troponin I (I for inhibition), along with tropomyosin, inhibits the interaction of actin and myosin by covering the myosin-binding site on actin. u Troponin C (C for Ca2+) is a Ca2+-binding protein that plays a central role in the initiation of contraction. When the intracellular Ca2+ concentration increases, Ca2+ binds to troponin C, producing a conformational change in the troponin complex. This conformational change moves tropomyosin out of the way, permitting the binding of actin to the myosin heads  Arrangement of Thick and Thin Filaments in Sarcomeres u The sarcomere is the basic contractile unit, and it is delineated 72 by the Z disks. u Each sarcomere contains a full A band in the center and one half of two I bands on either side of the A band. u The A bands are located in the center of the sarcomere and contain the thick (myosin) filaments, which appear dark when viewed under polarized light. u Thick and thin filaments may overlap in the A band; these areas of overlap are potential sites of cross-bridge formation. Arrangement of thick and thin filaments of skeletal muscle in sarcomeres 73 u The I bands are located on either side of the A band and appear light when viewed under polarized light. They contain the thin (actin) 74 filaments, intermediate filamentous proteins, and Z disks. They have no thick filaments. u The Z disks are darkly staining structures that run down the middle of each I band, delineating the ends of each sarcomere. u The bare zone is located in the center of each sarcomere. There are no thin filaments in the bare zone; thus, there can be no overlap of thick and thin filaments or cross-bridge formation in this region. u The M line bisects the bare zone and contains darkly staining proteins that link the central portions of the thick filaments together.  Cytoskeletal Proteins establish the architecture of the myofibrils, ensuring that the thick and thin filaments are 75 aligned correctly and at proper distances with respect to each other.  Transverse cytoskeletal proteins link thick and thin filaments, forming a "scaffold" for the myofibrils and linking sarcomeres of adjacent myofibrils. i. A system of intermediate filaments holds the myofibrils together, side by side. ii. The entire myofibrillar array is anchored to the cell membrane by an actin-binding protein called dystrophin. (In patients with muscular dystrophy, dystrophin is defective or absent.) u Longitudinal cytoskeletal proteins include two large proteins called titin and nebulin. 76 u Titin, which is associated with thick filaments, is a large molecular weight protein that extends from the M lines to the Z disks. u Part of the titin molecule passes through the thick filament; the rest of the molecule, which is elastic or springlike, is anchored to the Z disk. u As the length of the sarcomere changes, so does the elastic portion of the titin molecule. u Titin also helps center the thick filaments in the sarcomere. u Nebulin is associated with thin filaments. A single nebulin molecule extends from one end of the thin filament to the other. Nebulin serves as a "molecular ruler," setting the length of thin filaments during their assembly. α-Actinin anchors the thin filaments to the Z disk. The transverse tubules are continuous with the sarcolemmal membrane and invaginate deep into 77 the muscle fiber, making contact with terminal cisternae of the sarcoplasmic reticulum. Transverse tubules and sarcoplasmic reticulum of skeletal muscle. u The sarcoplasmic reticulum is an internal tubular structure, which is the site of storage and release of Ca2+ for excitation-contraction coupling. 78 u Its contains a Ca2+-release channel called the ryanodine receptor u Ca2+ is accumulated in the sarcoplasmic reticulum by the action of Ca2+ ATPase (SERCA) in the sarcoplasmic reticulum membrane. u This ensure that the intracellular Ca2+ concentration is kept low when the muscle fiber is at rest. u Within the sarcoplasmic reticulum, Ca2+ is bound to calsequestrin thus helps to maintain a low free Ca2+ concentration inside the sarcoplasmic reticulum, thereby reducing the work of the Ca2+ ATPase pump. u Thus, a large quantity of Ca2+ can be stored inside the sarcoplasmic reticulum in bound form, while the intrasarcoplasmic reticulum free Ca2+ concentration remains extremely low. u EXCITATION-CONTRACTION COUPLING IN SKELETAL MUSCLE u The mechanism that translates the muscle action potential 79 into the production of tension is excitation-contraction coupling. The steps involved in excitation-contraction coupling 1. Action potentials in the muscle cell membrane are propagated to the T tubules by the spread of local currents. 2. Depolarization of the T tubules causes a critical conformational change in its voltage-sensitive dihydropyridine receptor. This conformational change opens the Ca2+-release channels (ryanodine receptors) on the nearby sarcoplasmic reticulum 3. Ca2+ is released from its storage site in the sarcoplasmic reticulum into the ICF of the muscle fiber, resulting in an increase in intracellular Ca2+ concentration. 4. Ca2+ binds to troponin C on the thin filaments, causing a conformational 80 change in the troponin complex. 5. The conformational change in troponin causes tropomyosin (which was previously blocking the interaction of actin and myosin) to be moved out of the way so that cross-bridge cycling can begin 6. Cross-bridge cycling. With Ca2+ bound to troponin C and tropomyosin moved out of the way, myosin heads can now bind to actin and form so-called cross- bridges. Formation of cross-bridges is associated with hydrolysis of ATP and generation of force. 7. Relaxation occurs when Ca2+ is reaccumulated in the sarcoplasmic reticulum by the Ca2+ ATPase of the sarcoplasmic reticulum membrane (SERCA). When the intracellular Ca2+ concentration decreases to less than 10-7 M, there is insufficient Ca2+ for binding to troponin C. When Ca2+ is released from troponin C, tropomyosin returns to its resting position, where it blocks the myosin- binding site on actin. As long as the intracellular Ca2+ is low, cross-bridge cycling cannot occur, and the muscle will be relaxed u MECHANISM OF TETANUS u A single action potential results in the release of a fixed amount of 81Ca2+ from the sarcoplasmic reticulum, which produces a single twitch. u The twitch is terminated (relaxation occurs) when the sarcoplasmic reticulum reaccumulates this Ca2+. u However, if the muscle is stimulated repeatedly, there is insufficient time for the sarcoplasmic reticulum to reaccumulate Ca2+, and the intracellular Ca2+ concentration never returns to the low levels that exist during relaxation. u Instead, the level of intracellular Ca2+ concentration remains high, resulting in continued binding of Ca2+ to troponin C and continued cross- bridge cycling. u In this state, there is a sustained contraction called tetanus, rather than just a single twitch.  LENGTH-TENSION RELATIONSHIP The length-tension relationship in muscle refers to the effect of muscle fiber length on the amount of tension the fiber can develop. 82 u The amount of tension is determined for a muscle undergoing an isometric contraction, in which the muscle is allowed to develop tension at a preset length (called preload) but is not allowed to shorten. (Imagine trying to lift a 500-pound barbell. The tension developed would be great, but no shortening or movement of muscle would occur!) The following measurements of tension can be made as a function of preset length (or preload): u Passive tension is the tension developed by simply stretching a muscle to different lengths. (Think of the tension produced in a rubber band as it is progressively stretched to longer lengths.) u Total tension is the tension developed when a muscle is stimulated to contract at different preloads. It is the sum of the active tension developed by the cross-bridge cycling in the sarcomeres and the passive tension caused by stretching the muscle. u Active tension is determined by subtracting the passive tension from the total tension. It represents the active force developed during cross-bridge cycling  83 FORCE-VELOCITY RELATIONSHIP u The force-velocity relationship is determined by allowing the muscle to shorten. The force is fixed, rather than the length, and therefore, it is called an isotonic contraction. As the afterload on the muscle increases, the velocity will be decreased because cross-bridges can cycle less rapidly against the higher resistance. As the afterload increases to even higher levels, the velocity of shortening is reduced to zero.  Smooth Muscle u Smooth muscle lacks striations, which distinguishes it from 84 skeletal and cardiac muscle. The striations found in skeletal and cardiac muscle are created by the banding patterns of thick and thin filaments in the sarcomeres. In smooth muscle, there are no striations because the thick and thin filaments, while present, are not organized in sarcomeres. u Smooth muscle is found in the walls of hollow organs, such as the gastrointestinal tract, the bladder, and the uterus, as well as in the vasculature, the ureters, the bronchioles, and the muscles of the eye. The functions of smooth muscle are twofold: to produce motility (e.g., to propel chyme along the gastrointestinal tract or to propel urine along the ureter) and to maintain tension (e.g., smooth muscle in the walls of blood vessels). u TYPES OF SMOOTH MUSCLE 85 u Smooth muscles are classified as multiunit or unitary, depending on whether the cells are electrically coupled. Unitary smooth muscle has gap junctions between cells, which allow for the fast spread of electrical activity throughout the organ, followed by a coordinated contraction. Multiunit smooth muscle has little or no coupling between cells. A third type, a combination of unitary and multiunit smooth muscle, is found in vascular smooth muscle. u Unitary (single unit) smooth muscle is present in the gastrointestinal tract, bladder, uterus, and ureter. The smooth muscle in these organs contracts in a coordinated fashion because the cells are linked by gap junctions. Gap junctions are low-resistance pathways for current flow, which permit electrical coupling between cells. For example, action potentials occur simultaneously in the smooth muscle cells of the bladder so that contraction (and emptying) of the entire organ can occur at once. u Unitary smooth muscle is also characterized by spontaneous pacemaker activity, or slow waves. The frequency of slow waves sets a characteristic pattern of action potentials within an organ, which then determines the frequency of contractions. u Multiunit smooth muscle is present in the iris, in the ciliary muscles of the lens, and in the vas deferens. Each muscle fiber behaves as a separate motor unit (similar to skeletal muscle), and there is little or no coupling between cells. Multiunit smooth muscle cells are densely innervated by postganglionic fibers of the parasympathetic and sympathetic nervous systems, and it is these innervations that regulate function. u EXCITATION-CONTRACTION COUPLING IN SMOOTH MUSCLE 86 u The mechanism of excitation-contraction coupling in smooth muscle differs from that of skeletal muscle. u Recall that in skeletal muscle binding of actin and myosin is permitted when Ca2+ binds troponin C. u In smooth muscle, however, there is no troponin. Rather, the interaction of actin and myosin is controlled by the binding of Ca2+ to another protein, calmodulin. In turn, Ca2+-calmodulin regulates myosin-light-chain kinase, which regulates cross-bridge cycling. The steps involved in excitation-contraction coupling in smooth muscle: u Action potentials occur in the smooth muscle cell membrane. The depolarization 87 of the action potential opens voltage-gated Ca2+ channels in the sarcolemmal membrane. With the Ca2+ channels open, Ca2+ flows into the cell down its electrochemical gradient. This influx of Ca2+ from the ECF causes an increase in intracellular Ca2+ concentration. u Two additional mechanisms may contribute to the increase in intracellular Ca2+ concentration: 1. Ligand-gated Ca2+ channels: Ligand-gated Ca2+ channels in the sarcolemmal membrane may be opened by various hormones and neurotransmitters, permitting the entry of additional Ca2+ from the ECF. 2. Inositol 1,4,5-triphosphate (IP3)-gated Ca2+ release channels: IP3-gated Ca2+ release channels in the membrane of the sarcoplasmic reticulum may be opened by hormones and neurotransmitters. Either of these mechanisms may augment the rise in intracellular Ca2+ concentration caused by depolarization. u The rise in intracellular Ca2+ concentration causes Ca2+ to bind to calmodulin. Like troponin C in skeletal muscle, calmodulin binds four ions of Ca2+ in a cooperative fashion. The Ca2+- 88 calmodulin complex binds to and activates myosin-light-chain kinase. u When activated, myosin-light-chain kinase phosphorylates myosin. When myosin is phosphorylated, it binds actin to form cross-bridges, producing contraction. When myosin is in this phosphorylated state, cross-bridges can form and break repeatedly. One molecule of ATP is consumed with each cross-bridge cycle. The amount of tension generated is directly proportional to the number of cross-bridges formed, which is, in turn, proportional to the intracellular Ca2+ concentration. u When the intracellular Ca2+ concentration decreases, myosin is dephosphorylated by myosin- light-chain phosphatase. In the dephosphorylated state, myosin can still interact with actin, but the attachments are called latch-bridges rather than cross-bridges. The latch-bridges do not detach, or they detach slowly; thus, they maintain a tonic level of tension in the smooth muscle with little consumption of ATP. u Relaxation occurs when the sarcoplasmic reticulum reaccumulates Ca2+, via the Ca2+ ATPase, and lowers the intracellular Ca2+ concentration below the level necessary to form Ca2+- calmodulin complexes. 89  Mechanisms That Increase Intracellular Ca2+ Concentration in Smooth Muscle u During the action potential in smooth muscle, Ca2+ enters the cell from ECF 90 via sarcolemmal voltage-gated Ca2+ channels, which are opened by depolarization. This is only one source of Ca2+ for contraction. u Ca2+ also can enter the cell through ligand-gated channels in the sarcolemmal membrane, or it can be released from the sarcoplasmic reticulum by IP3-gated mechanisms. u In contrast, in skeletal muscle the rise in intracellular Ca2+ concentration is caused exclusively by depolarization-induced release from the sarcoplasmic reticulum- Ca2+ does not enter the cell from the ECF. u The three mechanisms involved in Ca2+ entry in smooth muscle are described as follows: u Voltage-gated Ca2+ channels are sarcolemmal Ca2+ channels that open when the cell membrane potential depolarizes. Thus, action potentials in the smooth muscle cell membrane cause voltage-gated Ca2+ channels to open, allowing Ca2+ to flow into the cell down its electrochemical potential gradient.  Ligand-gated Ca2+ channels also are present in the sarcolemmal membrane. 91 u They are not regulated by changes in membrane potential, but by receptor-mediated events. u Various hormones and neurotransmitters interact with specific receptors in the sarcolemmal membrane, which are coupled via a GTP-binding protein (G protein) to the Ca2+ channels. u When the channel is open, Ca2+ flows into the cell down its electrochemical gradient. u IP3-gated sarcoplasmic reticulum Ca2+ channels also are opened by hormones and neurotransmitters. 92 u The process begins at the cell membrane, but the source of the Ca2+ is the sarcoplasmic reticulum rather than the ECF. u Hormones or neurotransmitters interact with specific receptors on the sarcolemmal membrane (e.g., norepinephrine with α1 receptors). These receptors are coupled, via a G protein, to phospholipase C (PLC). u Phospholipase C catalyzes the hydrolysis of phosphatidylinositol 4,5- diphosphate (PIP2) to IP3 and diacylglycerol (DAG). u IP3 then diffuses to the sarcoplasmic reticulum, where it opens Ca2+ release channels (similar to the mechanism of the ryanodine receptor in skeletal muscle). u When these Ca2+ channels are open, Ca2+ flows from its storage site in the sarcoplasmic reticulum into the ICF.  93 Mechanisms for increasing intracellular [Ca2+] in smooth muscle. success 94