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As far aas I understand, in mitochondria, the citric acid cycle breaks down fatty acid or glucose to produce NADH and FADH2, which are then utilized by Complexes I through IV to generate a proton gradient in the intermembrane space. This gradient powers ATP synthase to ultimately generate ATP for the cell.

Uncoupling refers to alternative ways of decreasing the proton gradient without involving ATP synthase, such as through transmembrane fatty acids or Uncoupling Proteins (UCPs).

On the one hand, I recall learning that uncoupling is considered a disease condition since it disrupts the 'pull-effect' where an increased need for ATP essentially drives the entire reaction chain from the back, and because it generally dissipates energy into non-productive work. For instance, a textbook I recently read stated that ketones promote uncoupling in cardiac muscle cells, which is detrimental for obese individuals as the heart's performance capacity is impaired due to efficiency loss.

On the other hand, it is widely accepted nowadays that physical activity leads to significant mitochondrial uncoupling. This is surprising to me since one would assume that during a biologically relevant process, such as the adaptation of a muscle to movement, the adaptation would typically be 'beneficial'. Multiple authors write sentences like 'this optimizes the mitochondria's ability to synthesize ATP.'

It is evident that mitochondria deal better with a high nutrient supply when uncoupled, as fewer superoxides and peroxides are produced. But during physical exertion, an excess supply of nutrients is probably not the main issue?

I'm looking for insights to guide my understanding. How could the uncoupling of the electron transport chain I-IV from ATP synthase potentially enhance the performance capability of mitochondria and by this means the performance capability of skeletal muscle cells?

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Skeletal muscles have multiple systems that increase the rate of energy generation in response to faster energy consumption during excercise.

System Rate Sustainability
Phosphagen highest 10-15s
Glycogen-lactic-acid 30-40s
Aerobic lowest indefinite

Phosphagen system The phosphagen system represents the immediate source of ATP, it is relied on for power surges. ATP itself and phosphocreatine, which can donate its phosphate radical to ADP, form this system.

When muscle activity is prolonged, the other two systems come into play. Which of the two being predominant depends on the intensity of the activity (rate of ATP utilisation). Lactic acid fermentation The glycogen-lactic-acid system is used for strenuous excercise because it generates ATP faster than the aerobic system. It involves glycogenolysis, glycolysis and homolactic fermentation. Glycogenolysis is the breakdown of glycogen to glucose. Glycolysis is glucose oxidation into pyruvic acid by oxidative coenzymes with production of ATP (substrate-level phosphorylation). In homolactic fermentation, which happens under anaerobic conditions (oxygen insufficiency), instead of the reduced coenzymes (byproducts of glycolysis) being oxidised by the ETC in the mitochonderia, pyruvic acid acts as the final hydrogen acceptor to oxidise them so they can be used for another cycle of glycolysis. This produces lactic acid which is a strong acid that causes fatigue as will be explained later. Krebs' cycle is not part of the glycogen-lactic-acid system because there is no pyruvic acid to oxidatively decarboxylate into the acetyl moiety of acetyl-CoA (active acetate). Krebs' cycle and ETC The aerobic system involves glycogenolysis, glycolysis, Krebs' cycle and the ETC. The Kreb cycle substrate is the active acetate radical that is oxidatively decarboxylated completely by oxidative coenzymes, with substrate-level phosphorylation. The ETC requires oxygen supply since oxygen is the final electron acceptor that drives the movement of hydrogen from the reduced coenzymes and its splitting to electrons and protons. (Oxygen is the second most electronegative element in the periodic table.) The movement of electrons (redox reactions) generates energy that is used to pump protons in the matrix across the inner mitochonderial membrane into the intermembrane space. This pumping continues until a proton back-pressure (proton motive force or proton gradient) is established. Once established, ATP synthase inhibits further proton pumping and stops the activity of the ETC. The pumped protons then diffuse through ATP synthase (chemiosmosis) and the energy that was used to pump them before is now used to phosphorylate ADP to ATP (oxidative phosphorylation). ATP synthase is sometimes also called complex V though it doesn't transport electrons, only facilitates proton diffusion. ATP synthase is activated by high ADP concentration in the matrix which discharges the proton back-pressure and consequently activates the ETC. ADP is produced from ATP hydrolysis and increases at low energy states when energy consumption is faster than energy production. The aerobic system is thus relied on for moderate-intensity prolonged excercise where oxygen consumption by the ETC is balanced by oxygen supply and where the long metabolic pathway still provides ATP at a sufficient rate. The reduction of oxygen by electrons can form either

  • water if the electrons are sufficient $\ce{O2 + 4e- + 4H+ \to 2H2O}$ (The protons are taken up from the matrix).
  • superoxide ion if the electrons are insufficient to form water $\ce{O2 + e- \to [O2]-}$.

Superoxide ions form hydrogen peroxide $\ce{2[O2]- + 2H+ \to H2O2 + O2}$. Hydrogen peroxide, as with all ROS, has a high affinity to electrons and pulls them from other compounds damaging them, to combine with the electrons and with protons forming water $\ce{H2O2 + 2e- + 2H+ \to 2H2O}$. This causes peroxidation (oxidative damage) to lipids, proteins and DNA. Cells therefore have to split hydrogen peroxide to water and oxygen $\ce{2H2O2 \to 2H2O + O2}$ by catalase enzyme. Hydrogen peroxide can also be useful for the same reason. It is used by immunity cells to kill pathogens by damaging their cell membranes, which are made up of phopholipids, causing their lysis.

The muscle burns through the phosphagen system first and then operates through the aerobic system indefinitely, as long as there is sufficient oxygen and glucose supply. Any extra energy needed in a short amount of time is supplied by the glycogen-lactic-acid system. However, the latter limits activity time (causes fatigue) as will be explained later.


Skeletal muscle structure Muscles are divided into fascicles which are groups of muscle fibers (cells). Following are the most important organelles in a muscle fiber.

  • A sarcolemma (plasmalemma)
  • A sarcoplasm (cytoplasm)
  • Many mitochonderia
  • Multiple nuclei
  • Many sarcoendoplasmic reticula (smooth endoplasmic reticula) that surround the most important organelles, myofibrils
  • Many myofibrils

Myofibrils are formed of many myofilaments (actin and myosin) organized into contractile (motor) units known as sarcomeres. Neuromuscular junction and triad Muscles contract when a threshold electrical stimulus depolarises the sarcolemma resulting in an action potential that propagates to T-tubules. At the T-tubules, the reversal of relative charge accross the membrane changes the shape of dihydropyridine receptors so they now allow for the influx of $\ce{Ca^{2+}}$. Consequently, ryanodine receptors on the terminal cisternae of sarcoendoplasmic reticula open and more $\ce{Ca^{2+}}$ diffuse into the sarcoplasm ($\ce{Ca^{2+}}$-induced $\ce{Ca^{2+}}$ release). This increases sarcoplasmic $\ce{Ca^{2+}}$ concentration. $\ce{Ca^{2+}}$ binds to troponin which changes shape. Since troponin is bound to tropomyosin, changing the shape of the former displaces the latter. Tropomyosin consequently exposes myosin-binding sites on actin so myosin can now bind, bend, detach and get back to original conformation to repeat the process (cross-bridge cycling) until $\ce{Ca^{2+}}$ is actively pumped back to where it came from causing tropomyosin to block active sites and relaxation. The magnitude of the response of the muscle to stimulation (whether acetylcholine or depolarisation can elicit an action potential) depends on

  • strength of the stimulus (amount of acetylcholine or change in membrane potential).
  • duration of the stimulus (time for which acetylcholine or charge persist).
  • frequency of stimulation or rise of stimulus intensity (since acetylcholine and charge accumulate).

Muscle fatigue as we know it can be attributed to different mechanisms that happen at different stations.

  • Neural (central) fatigue
  • Metabolic (peripheral) fatigue

Neural fatigue happens because the muscle stops listening, specifically because the frequency of electric stimulation is not high enough to maintain contraction. It can happen because of exhaustion of neurotransmitter (acetylcholine) stores for example. That is to say, the rate of release and breakdown of acetylcholine by acetylcholinesterase is greater than the rate of its formation and packaging into vesicles. Neural fatigue is what limits activity time in well-trained athletes. It's not associated with pain. Athletes work on increasing the capacity of their neurons to keep firing at a high frequency by training.

Metabolic fatigue on the other hand happens because the muscle gives up, specifically because of

  • $\ce{Ca^{2+}}$ not doing its job being too little or ineffective.
  • substrate shortage.

Lactic acid, being highly acidic, dissociates readily into protons and lactate. The protons, having the same charge as $\ce{Ca^{2+}}$, displace it decreasing the sensitivity of the muscle to $\ce{Ca^{2+}}$. The binding of protons can also deform proteins by breaking the bonds that form their secondary, tertiary and quaternary structures. Amino acid radical groups pick up the surplus of positive charge and the interactions between them change. Thus, lactic acid causes inhibition of active $\ce{Ca^{2+}}$ pumping outside the sarcoplasm by deforming pumps, so high $\ce{Ca^{2+}}$ concentration is maintained allowing for more forceful contraction. However, it's a matter of time until the insensitivity becomes too much. Lactic acid (which would have been an active acetate inside mitochonderia in the aerobic system) also causes hydrolytic damage to myofilaments. Any damage to a muscle is called microtrauma. Repetitive strain on myofilaments, connective tissue, tendons and bones by lengthening (eccentric contraction) can also cause microtrauma. Microtrauma results in inflammation and muscle soreness. Muscle soreness (pain) aims to stop the physical activity which can cause more microtrauma to the muscle. Muscles can adapt to metabolic fatigue and microtrauma by increasing

  • substrate stores (glycogen)
  • $\ce{Ca^{2+}}$ stores (sarcoendoplasmic reticula)
  • number of myofibrils (hypertrophy)
  • number of connective tissue fibers, to increase stiffness
  • diameter of nerve fiber, to increase speed of conduction
  • amount of glycolytic enzymes

as seen in pale (also known as type II or fast-twitch) muscle fibers, to decrease the likelihood of reinjury.


Thus, muscles don't necessarily need the ETC to generate energy and in fact, rely on the fermentation pathway for faster and more forceful activities because of its higher rate of ATP production despite its fatiguing effects which they can adapt to.


Now, we can look at the effects of uncoupling. Uncouplers work on the aerobic pathway by uncoupling oxidation from phosphorylation; they make it possible for oxidation to take place without phosphorylation. They cause proton discharge through the inner mitochonderial membrane before the threshold back-pressure required for protons to flow through ATP synthase is established. As a result, the energy from proton diffusion across the membrane is released as heat instead. This mechanism is utilized for example in brown adipose tissue where thermogenin transmembrane protein (uncoupling protein 1) causes rising of body temperature (to maintain body temperature in cold environments) and fat loss by metabolising fatty acids in stored fat which enter Krebs' cycle as acetyl groups and the resulting reduced coenzymes are oxidised by the ETC, with no oxidative phosphorylation. That's why this type of adipose tissue is more vascular relative to white adipose tissue; to maintain the high rate of oxidation which requires oxygen. The increase in heat can

  • decrease viscosity of myofilaments so they slide more easily over each other.
  • increase enzyme activity since more enzyme-substrate complexes form as enzymes and substrates move faster with greater chances to collide which increases the rate of ATP formation and utilisation.
  • relax blood vessel smooth muscles resulting in vasodilatation and increased blood flow, with consequent increases in oxygen supply and lactic acid removal during excercise. Whether lactic acid causes fatigue depends on the difference between the rate of its production and the rate of its flushing during activity. At rest, the remaining lactic acid is flushed or oxidised locally. We maintain the vasodilatation and high ventillation rate at rest, to oxidise the remaining lactic acid and myoglobin and form ATP by the ETC to rephosphorylate creatine to phosphocreatine (oxygen debt). The flushed lactic acid is mostly reoxidised into pyruvic acid then glucose in the liver by reversal of glycolysis, also known as gluconeogenesis (Cori cycle). The remaining lactic acid is reoxidized locally to pyruvic acid which enters Krebs' cycle instead, but the majority is flushed. The increased blood supply also helps in healing from microtrauma at rest which ultimately stops soreness. Cold, on the other hand, treats soreness directly by decreasing blood flow carrying inflammatory cells that cause pain by secreting cytokines.

Besides increasing temperature, uncoupling causes

  • faster flow of electrons through the ETC since the sufficient proton back-pressure is never established so ATP synthase doesn't inhibit pumping of protons and electron transfer (electron transfer is continuous). As a result, oxygen is reduced rapidly to water instead of being reduced merely to hydrogen peroxide that can cause damage if catalase is saturated and can't split the excess to water and oxygen.
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    $\begingroup$ Wow! What did you do? That's 1/2 a textbook! 1000s kudos! Great job! I really appreciate your effort! $\endgroup$
    – DrSvanHay
    Jul 30 at 15:44
  • $\begingroup$ Thank you. I truly appreciate your nice words. It just so happens I am studying biochemistry and physiology. So I thought why not try writing a logical, comprehensive, self-contained answer that makes total sense to make sure I understand what I am learning that can also be my reference since it integrates everything. My former answer also didn't address peroxidation so I had to fix it. $\endgroup$ Jul 30 at 17:48

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