AnSc310-Lecture 2. Movement of Substances Across Membranes - Physiology Notes

Structure and General Functions of the Cell Membrane

Phospholipid Bilayer Structure

• The cell membrane is organized as a amphipathic phospholipid bilayer consisting of two lipid layers:
  • Glycerol backbone: Forms the structural base of each phospholipid molecule.
  • Two fatty acid chains: Form hydrophobic (water-fearing) nonpolar tails oriented toward the interior of the membrane, away from water.
  • Phosphate group: Forms a hydrophilic (water-loving) polar head facing outward toward the aqueous intracellular fluid (ICF) and extracellular fluid (ECF).

Lipid and Protein Composition

• Cell membranes are primarily composed of three main lipid types:
  1. Phospholipids: Main structural bilayer component.
  2. Sphingolipids: Specialized structural membrane lipids.
  3. Cholesterol: Interspersed between phospholipids to regulate membrane fluidity and mechanical stability.
• Associated membrane proteins and carbohydrates include:
  • Integral proteins: Embedded deeply within or spanning across the lipid bilayer.
  • Channel proteins: Transmembrane proteins providing aqueous pores across the membrane.
  • Glycoproteins: Membrane proteins covalently conjugated with carbohydrates (acting as recognition sites and receptors).
  • Glycolipids: Membrane lipids conjugated with carbohydrate chains.

General Functions of Cell Membranes

1. Physical isolation:
   • Serves as a physical barrier separating the intracellular fluid (ICF) from the extracellular fluid (ECF).
2. Regulation of exchange with the environment:
   • Regulates the controlled entry of essential ions and nutrients into the cell.
   • Facilitates the elimination of metabolic waste products (e.g., urea produced from metabolic degradation, which is transported out into the blood for renal excretion).
   • Governs the release and secretion of cell-synthesized products (e.g., hormones, digestive enzymes).
3. Communication between the cell and its environment:
   • Contains membrane receptor proteins that recognize specific extracellular signal molecules (ligands such as hormones or neurotransmitters) and trigger specific biological responses inside the cell.
4. Structural support:
   • Membrane proteins anchor cytoskeletal elements, maintaining cell shape, structural integrity, and positioning cellular organelles.

Selective Permeability

• Cell membranes are semipermeable and selectively permeable: they permit specific substances to cross while completely blocking others.
• Permeable substances: Nonpolar molecules and small polar molecules dissolve directly in the lipid bilayer and cross without protein assistance (e.g., $O_2$, $CO_2$, fatty acids, $H_2O$).
• Impermeable substances: Ions and large polar molecules cannot cross the lipid bilayer directly and require protein carriers or channels (e.g., glucose, proteins, $Na^+$, $K^+$, $Ca^{2+}$, $Cl^-$).
• Physiological example: The blood-brain barrier (BBB) enforces strict selective permeability to isolate central nervous system neurons from harmful systemic circulating substances, pathogens, and metabolic fluctuations.

Compositional Differences Between Intracellular and Extracellular Fluids

• Potassium ($K^+$): Highly concentrated inside the cell (ICF $\neq 140\,mM$ or $\text{mmol/L}$) compared to outside (ECF $\neq 4\,mM$).
• Sodium ($Na^+$): Highly concentrated outside the cell (ECF $\neq 145\,mM$) compared to inside (ICF $\neq 5\,mM$).
• Proteins and Amino Acids: Highly concentrated inside the cell (ICF); negatively charged intracellular amino acids and proteins impart a net negative charge to the cell interior.
• Adenosine Triphosphate ($ATP$): Present exclusively inside the cell (ICF) as an intracellular energy source generated by mitochondrial glucose and nutrient degradation; $ATP$ does not exist in ECF and does not leak across healthy membranes.

Classification of Transport Mechanisms

Passive Transport

• Definition: Transport of substances across membranes occurring naturally without cellular metabolic energy expenditure.
• Key Characteristics:
  • Spontaneous process driven by inherent thermal energy (Brownian motion) of particles.
  • Does not require $ATP$ or cellular metabolic energy.
  • Solute movement is "downhill" (following concentration, electrical, or electrochemical gradients).
• Major Subdivisions:
  1. Simple diffusion.
  2. Channel-mediated diffusion.
  3. Facilitated diffusion.

Active Transport

• Definition: Transport of substances across membranes mediated by membrane proteins requiring energy input.
• Key Characteristics:
  • Non-spontaneous process requiring cellular energy, typically from $ATP$ hydrolysis.
  • Solute movement is "uphill" (against concentration, electrical, or electrochemical gradients).
  • Highly regulated by cellular control mechanisms and membrane transporter proteins.
• Major Subdivisions:
  1. Primary active transport.
  2. Secondary active transport.

Factors Affecting the Direction of Transport: Driving Forces

Terminology and Fundamentals

• Driving Force: Energy acting upon particles that governs their movement direction and rate.
• Push: Force acting in the direction of motion, supporting or accelerating particle movement.
• Pull: Force acting against the direction of motion, slowing or stopping particle movement.
• Particle: Sub-microscopic unit of matter, including atoms, ions, and molecules.

Chemical Driving Force

• Concentration Gradient ($\Delta C$): Difference in particle concentration (number of molecules per unit volume) between two adjacent regions.
• Mechanism: Driven by random, chaotic thermal motion of particles (Brownian motion). Particles do not possess intent; random collisions cause a net statistical movement from crowded regions to less crowded regions.
• Direction: Always acts down the concentration gradient, moving particles from higher concentration to lower concentration.

• Net Flux:
  • Gross particle movement occurs in both directions across a permeable boundary.
  • Net flux represents the directional difference in particle transport per unit time.
  • At concentration equilibrium ($\Delta C = 0$), particle movement continues equally in both directions, making net flux equal to zero ($\text{Net Flux} = 0$).
• Magnitude: The magnitude of the chemical driving force is directly proportional to the concentration gradient ($\Delta C$). A larger concentration gradient produces a stronger chemical driving force.

Electrical Driving Force

• Ions: Charged atoms or molecules.
  • Cation: Positively charged ion (e.g., $Na^+$, $K^+$, $Ca^{2+}$).
  • Anion: Negatively charged ion (e.g., $Cl^-$, $HCO_3^-$).

• Membrane Potential ($V_m$):
  • The electrical potential difference (voltage) existing across the plasma membrane, generated by an unequal distribution of cations and anions across the membrane.
  • Under normal resting physiological conditions, there is a slight excess of anions in the ICF clustered along the inner membrane surface, and a slight excess of cations in the ECF clustered along the outer membrane surface.
  • This charge separation acts like a microscopic cellular battery storing potential energy.

• Measurement and Polarity Convention:
  • Measured in millivolts ($mV$), where $1\,mV = 10^{-3}\,V$.
  • Polarity convention defines $V_m$ relative to the inside of the cell (ICF).
  • Typical resting membrane potential in many excitable cells is $V_m \approx -70\,mV$ (and ranges to $V_m = -85\,mV$ in skeletal muscle cells).
• Electrostatic Principles:
  • Attraction of opposite charges: Opposite charges attract each other (e.g., positive $Na^+$ attracted to negative $Cl^-$).
  • Repulsion of like charges: Like charges repel each other (e.g., $Na^+$ repels $Ca^{2+}$; $Cl^-$ repels $HCO_3^-$).
• Direction of Electrical Driving Force:
  • Cations (+): Pulled inward toward the negatively charged intracellular environment.
  • Anions (-): Pushed outward away from the negatively charged intracellular environment toward the positively charged extracellular fluid.
• Magnitude Factors of Electrical Driving Force:
  1. Strength of Membrane Potential ($V_m$): A larger absolute membrane voltage difference exerts a stronger electrical pull or push on charged ions (e.g., $V_m = -100\,mV$ exerts a stronger inward force on cations than $V_m = -50\,mV$).
  2. Ion Valency ($z$): The quantity of electrical charge on the ion. At an identical $V_m$, a divalent ion ($z = \pm 2$, e.g., $Ca^{2+}$) experiences twice the electrical driving force of a monovalent ion ($z = \pm 1$, e.g., $K^+$).

• Physiological Importance of Membrane Potential:
  • Nerve signals: Essential for generating and propagating neuronal action potentials.
  • Muscle contraction: Triggers mechanical contraction in skeletal, cardiac, and smooth muscle fibers.
  • Transport energy: Powers secondary transport processes for nutrients and ions.
  • Synaptic transmission: Voltage changes trigger localized $Ca^{2+}$ entry at presynaptic terminals, causing neurotransmitter vesicle exocytosis.

Electrochemical Driving Force (ECDF)

• Definition: The net combined force acting on an ion, representing the vector sum of both the chemical driving force and the electrical driving force.

• Case 1: Forces Act in the Same Direction:
  • Chemical force and electrical force reinforce one another.
  • Magnitude of ECDF equals the sum of both forces: $\text{ECDF} = F_{\text{chem}} + F_{\text{elec}}$.
  • Results in a stronger net force and rapid ion movement in that direction.
• Case 2: Forces Act in Opposite Directions:
  • Chemical force and electrical force oppose one another.
  • Net ion transport moves in the direction of whichever force is stronger.
  • Magnitude of ECDF equals the numerical difference between the stronger and weaker force: $\text{ECDF} = |F_{\text{chem}} - F_{\text{elec}}|$.
  • Ion transport occurs in the direction of the dominant force, but at a reduced net rate.

Equilibrium Potential ($E_x$)

• Definition: The precise membrane potential ($V_m$) at which the electrical driving force acting on ion $X$ exactly equals and balances the chemical driving force acting on ion $X$.
• Key Takeaway: At equilibrium potential ($V_m = E_x$), the net electrochemical driving force is zero ($\text{ECDF} = 0$), resulting in no net movement ($\text{Net Flux} = 0$) of that specific ion across the membrane.

• Potassium Equilibrium Potential ($E_K$) Example:
  • For potassium ($K^+$), the equilibrium potential is $E_K = -94\,mV$.
  • Chemical force acts outward (due to high intracellular $K^+$ concentration of $140\,mM$ vs $4\,mM$ extracellular).
  • Electrical force acts inward (due to negative intracellular membrane charge pulling positive $K^+$ ions inward).
  • When $V_m = E_K = -94\,mV$, the inward electrical pull exactly equals the outward chemical push.
• Comparison of $V_m$ and $E_K$ Governing Potassium Movement:
  1. $V_m$ is weaker (less negative) than $E_K$ (e.g., $V_m = -70\,mV$ vs $E_K = -94\,mV$):
  • At $-70\,mV$, the cell interior is insufficiently negative to hold $K^+$ inside electrically.
  • Outward Chemical Force > Inward Electrical Force.
  • Net Electrochemical Force is OUTWARD; $K^+$ exits the cell.
  1. $V_m$ is stronger (more negative) than $E_K$ (e.g., $V_m = -100\,mV$ vs $E_K = -94\,mV$):
  • At $-100\,mV$, the strong internal negativity pulls $K^+$ inward more powerfully than the chemical gradient pushes it out.
  • Inward Electrical Force > Outward Chemical Force.
  • Net Electrochemical Force is INWARD; $K^+$ enters the cell.
  1. $V_m = E_K = -94\,mV$:
  • Electrical Force = Chemical Force.
  • Net Electrochemical Force is ZERO; no net $K^+$ transport.

Passive Transport Mechanisms

Simple Diffusion

• Definition: Direct passive transport of small, nonpolar, uncharged, or lipid-soluble molecules directly through the phospholipid bilayer from high to low concentration.
• Characteristics:
  • Does not require membrane transport proteins or $ATP$.
  • Driven by random thermal energy motion.
  • Non-saturable linear rate kinetics: diffusion rate increases indefinitely in direct linear proportion to concentration increases.
• Five Key Factors Influencing Simple Diffusion Rate:
  1. Concentration gradient ($\Delta C$): Greater concentration difference across the membrane produces faster diffusion.
  2. Membrane surface area ($A$): Larger membrane surface area available increases total diffusion rate.
  3. Membrane permeability / Lipid solubility ($P$): Highly lipid-soluble or permeable molecules cross hydrophobic core faster.
  4. Molecular size / Molecular weight ($MW$): Smaller molecules diffuse faster than larger molecules.
  5. Pressure difference ($\Delta P$): Greater hydrostatic or partial pressure difference across the membrane accelerates diffusion rate.

• Veterinary Applications:
  • Respiratory gas exchange ($O_2$ and $CO_2$) across bovine, equine, or canine pulmonary alveoli and capillary membranes, and across fish gills.
  • Intestinal absorption of lipophilic nutrients and volatile fatty acids (VFAs) across rumen epithelium.
  • Renal tubular reabsorption of urea and uncharged solutes.

Channel-Mediated Diffusion

• Definition: Passive transport of ions or water across membranes through specialized transmembrane channel proteins down concentration or electrochemical gradients.
• Channel Protein Architecture:
  • Transmembrane proteins spanning the bilayer with a central hydrophilic pore (allowing polar ions or water to pass) surrounded by hydrophobic outer domains anchoring the channel within the lipid bilayer.

• Types of Channels:
  1. Aquaporins: Specialized transmembrane water channels mediating high-speed osmotic water movement (e.g., renal tubular fluid reabsorption).
  2. Ion Channels: Selective protein channels containing a selectivity filter permitting passage of specific ions ($Na^+$, $K^+$, $Cl^-$).
  3. Leak Channels: Continuously open channels permitting ongoing baseline passive ion flow (e.g., resting $K^+$ leak channels).
  4. Gated Channels: Channels possessing molecular gates that open or close in response to specific physical or chemical signals:
  • Voltage-gated channels: Open or close in response to alterations in membrane potential ($V_m$) (e.g., voltage-gated $Na^+$ and $K^+$ channels during neuronal action potentials).
  • Ligand-gated channels: Open or close upon binding specific chemical messengers (neurotransmitters, hormones).
  • Mechanically-gated channels: Open or close in response to physical stretch or membrane deformation.

Facilitated Diffusion (Carrier-Mediated Passive Transport)

• Definition: Passive movement of polar or larger molecules (e.g., glucose, amino acids) down their concentration gradient via specific transmembrane carrier proteins without energy ($ATP$) input.
• Mechanism of Glucose Transport via GLUT Carries:
  1. Carrier protein (GLUT) faces the extracellular space open to high solute concentration.
  2. Solute (glucose) binds specifically to a fitting binding site on the carrier.
  3. Binding induces a conformational change (shape shift) in the carrier protein.
  4. The carrier opens to the intracellular cytosol, reducing its binding affinity, and releases glucose into the cell.
  5. The carrier resets to its original outward-facing conformation.

• Saturable Kinetics ($V_{\text{max}}$):
  • Unlike simple diffusion, facilitated diffusion exhibits hyperbolic, saturable transport kinetics.
  • At low substrate concentrations, transport rate rises rapidly.
  • As substrate concentration increases, carrier binding sites become progressively occupied until reaching a maximal transport rate ($V_{\text{max}}$).
  • $V_{\text{max}}$ represents the maximum rate of facilitated diffusion occurring when 100% of available carrier proteins are saturated with substrate.

• Factors Affecting Transport Rate:
  1. Transport rate/turnover speed of individual carrier proteins.
  2. Abundance/density of carrier proteins in the membrane (upregulated via gene expression, hormones, or physical adaptation; e.g., exercise increases GLUT4 carrier density in equine muscle cells; lactation upregulates nutrient carriers in canine mammary cells and swine small intestine).
  3. Steepness of solute concentration gradient.
  4. Carrier saturation state.

Active Transport Mechanisms

Key Features

• Non-spontaneous process requiring cellular energy ($ATP$).
• Transports solutes "uphill" against concentration or electrochemical gradients (from low concentration to high concentration).
• Mediated by specialized protein pumps that function both as transporters and enzymes (ATPases that hydrolyze $ATP \rightarrow ADP + P_i$).
• Substrate-specific and saturable ($V_{\text{max}}$ kinetics).

Primary Active Transport

• Definition: Active transport in which energy released from direct $ATP$ hydrolysis is directly coupled to move solutes against their electrochemical gradient.

• Primary Example: The Sodium-Potassium Pump ($Na^+/K^+$-ATPase):

  • Stoichiometry and Sequential Steps per Cycle:
  1. Three $Na^+$ ions bind to high-affinity intracellular sites on the pump.
  2. $ATP$ is hydrolyzed ($\text{ATP} \rightarrow \text{ADP} + P_i$), transferring a phosphate group to phosphorylate the pump protein.
  3. Phosphorylation induces a conformational change that opens the pump to the ECF, lowers $Na^+$ affinity, and releases three $Na^+$ ions outside the cell.
  4. Two $K^+$ ions bind to high-affinity extracellular sites on the pump.
  5. Phosphate ($P_i$) is released, dephosphorylating the pump.
  6. Dephosphorylation returns the pump to its original intracellular-facing shape, lowers $K^+$ affinity, and releases two $K^+$ ions into the cytosol.
  • Electrogenic Property:
  • Per cycle, the pump extrudes three positive charges ($3\,Na^+$) while bringing in only two positive charges ($2\,K^+$).
  • This net removal of one positive charge per cycle makes the $Na^+/K^+$-ATPase electrogenic, directly maintaining the negative resting membrane potential.
  • Physiological Importance:
  • Maintains high extracellular $Na^+$ ($145\,mM$) and low intracellular $Na^+$ ($5\,mM$).
  • Maintains high intracellular $K^+$ ($140\,mM$) and low extracellular $K^+$ ($4\,mM$).
  • Crucial for osmotic volume control, cell excitability, and powering secondary active transport.

Secondary Active Transport

• Definition: Active transport mechanisms that do not hydrolyze $ATP$ directly, but instead utilize energy stored in ion concentration gradients previously established by primary active transport (primarily the steep inward $Na^+$ gradient created by the $Na^+/K^+$-ATPase).
• Types and Examples:
  1. Symport (Cotransport):
  • Driven ion and transported solute move in the SAME direction across the membrane.
  • Example: $Na^+$/Glucose Symporter (SGLT).
    • $Na^+$ diffuses down its steep electrochemical gradient into the cell, powering the simultaneous uptake of glucose against its concentration gradient into intestinal enterocytes or renal proximal tubule cells.
  1. Antiport (Counter-transport or Exchange):
  • Driven ion and transported solute move in OPPOSITE directions across the membrane.
  • Example 1: $Na^+/H^+$ Exchanger (NHE).
    • $Na^+$ moves into the cell down its gradient, driving the outward secretion of $H^+$ against its gradient to regulate intracellular pH.
  • Example 2: $Na^+/Ca^{2+}$ Exchanger (NCX).
    • $Na^+$ moves into the cell down its gradient, driving the extrusion of $Ca^{2+}$ against its gradient to maintain low cytosolic $Ca^{2+}$ levels required for cellular signaling.

Active-Passive Coordination and Dynamic Steady State

Dynamic Steady State vs. Thermodynamic Equilibrium

• Thermodynamic Equilibrium: Passive state where opposite fluxes are equal without energy expenditure ($\text{Net Flux} = 0$).
• Dynamic Steady State: Non-equilibrium state where continuous metabolic energy expenditure ($ATP$) maintains constant intracellular conditions over time despite continuous passive leakage.

The Pump-and-Leak Mechanism

• Active Pumping: The $Na^+/K^+$-ATPase actively transports $3\,Na^+$ out of the cell and $2\,K^+$ into the cell continuously using $ATP$.
• Passive Leaking: Passive leak channels permit continuous downhill leaking of $Na^+$ back into the cell and $K^+$ out of the cell down their electrochemical gradients.
• Steady State Equilibrium: Achieved when pump-mediated active flux exactly balances leak-mediated passive flux ($\text{Flux}_{\text{pump}} = \text{Flux}_{\text{leak}}$).
• This dynamic active-passive interplay stabilizes intracellular and extracellular ion concentrations, maintaining resting membrane potential, cell volume, and viability.