Introduction to Membrane Potentials Joseph G. Griswold, Ph.D. Dept. of Biology, The City College of New York I. Introduction In living things, most cell membranes have membrane potentials. This difference in electrical charge between the inside and outside of the membrane is the basis for many types of physiological processes, including transport of particles across the membrane and signaling among cells. It is estimated that in some cells up to 40% of energy is used to power active transport, a process that maintains or restores membrane potentials. The goal of this module is to enable you to understand the mechanisms through which potentials are developed and changed over time. II. Student Background To complete this module successfully, you should have or be provided with the following background: A. Basic operating skills for computers: open programs, navigate using a mouse, type in responses, etc. B. Definitions of chemical and electrical gradients. C. Graphing basics: reading axes, plotting points, stating in words the trends shown by a graph. D. Skills to read and follow simple written directions. III. Benchmarks for the Module Having completed this module, you will be able to: A. To represent a cell membrane by drawing a labeled diagram. B. To identifying the hydrophobic and hydrophilic elements, gated and passive channels, molecular pumps and the distribution of ions and molecules on the in- and outside of the membrane. C. To determine from a diagram the net charge in and outside the cell membrane using simple counting methods, and calculate the membrane potential in a static system using algebraic methods or the line method introduced in the module. D. To predict how ions will move through passive ion channels, given a starting ion distribution across the membrane. E. To represent a membrane potential on a graph and plot changes in potential over time from a data set that is provided. F. To state in words, using the terms resting membrane potential, depolarize, and hyperpolarize, the changes in membrane potential over time illustrated in a graph. G. To predict what will happen to membrane potential with the opening & closing of gated channels. H. To illustrate the opening of channels, flow of ions, and change in potential using a paper model. I. To graph changes in membrane potential for local potentials (PSPs) that depolarize and hyperpolarize the membrane and relate the segments of the curve to events in the membrane. J. To predict what happens to Na+ ions that flow into a neuron in producing local currents. IV. Prelab Activities A. The Intracellular and Extracellular Environments __1. Use class notes and text readings, as assigned by your instructor, to complete the following steps. We encourage you to work with a classmate. When you complete a step, check it off in the space beside the number. __2. The chemical makeup of cytoplasm inside a cell is quite different from that of the interstitial fluid (ISF) that bathes it (Table 1.1). It is with the ISF that cells directly exchange molecules and ions on a continuing basis. What are some molecules that are exchanged? _______________________________________________ . Table 1.1. Chemical makeup of the body fluids Substance Interstitial Fluid Intracellular Fluid Concentration Concentration Na+ 142 mEq/liter 10 mEq/liter K+ 4 140 Ca2+ 2.4 0.0001 Mg2+ 1.2 58.0 Cl- 103 4 HCO3- 28 10 Phosphate ion 28 10 Glucose 90-100 mg% 0-20 mg% Amino acids 4 75 Proteins 20k 160k pH 7.4 7.0 __3. Figure 1.1 is a simple diagram of 3 fluid compartments in the body. A capillary (A) is drawn to represent the blood or plasma compartment, which exchanges water and solutes directly with the interstitial fluid compartment (C). The intracellular "compartment" (B) is actually the millions of individual cytoplasmic spaces inside each cell of the body. Label the compartments in Fig. 1.1. __4. Walls of most capillaries are quite leaky, and the gaps allow free exchange of water, small molecules and ions between the plasma and interstitial fluid. In Figure 1.1, draw a small circle to represent an oxygen molecule (O2) in the blood, then use an arrow to indicate the direction of movement between the blood and the cell. Do the same for CO2. Some large particles like protein molecules in the blood cannot normally cross the capillary wall, nor can most of the blood cells. __5. Regulation of molecular exchange between the ISF and the intracellular compartments is accomplished by the cell membrane. You may have learned that the cell membrane is semi- permeable. What does that term mean to you? ____________________________________ ______________________________________________________________________ . __ 6. Figure 1.2 represents a piece of the cell membrane with the interstitial fluid above and the cytoplasm below. Label the compartments. __ 7. You may remember that an important part of the cell membrane is the phospholipid bilayer that is represented in this figure. Each phospholipid molecule in the bilayer has a phosphate head that is hydrophilic (attracted to water), and a pair of fatty acid tails that are hydrophobic (water repelling). The arrangement of the bilayer occurs automatically as a consequence of how each end of the molecule interacts with water. Looking at the diagram, can you briefly explain how this happens? _____________________________________________ _______________________________________________________________________ . __ 8. Missing in Fig 1.2 are cholesterol molecules and the large proteins that are usually shown embedded in the cell membranes. We will study more about these important elements later. B. Concentrations, Gradients, and Permeability. __1. You may remember that in all fluid compartments of the body, solute molecules or ions (for example sodium ions or glucose) are dissolved in the solvent water. At physiological temperatures, the solute particles move rapidly (Brownian motion), colliding with one another and with the membranes of the cell. __ 2. Also, each solute is present in a particular concentration. To review: Concentration refers to the number of solute particles per unit volume of solution. There are several measures, two of which are given in Table 1.1. What are they? _______________________________________ . __ 3. Look at Figure 1.2 again. Assume for our work that the volume of solution both inside and outside the membrane is 0.1 ml. What would be the concentration of small molecules outside in the interstitial fluid? Use the measure "particles/ml" as the unit of measure. Enter the results into the data box. Concentration __ 4. What are the concentrations of medium sized molecules both in- and outside the cell? Inside Outside __ 5. Concentration gradients exist in situations where the number of molecules per unit volume (concentration) at one location differs from that at another. In the prior item, you discovered that a concentration gradient exists across the cell membrane. The "high" end of the gradient is the location where the concentration is greatest. The other is the "low" end. In diffusion, the net movement of molecules by random motion is down the concentration gradient, from the high to the low end. Do molecules in a gradient ever diffuse up the gradient? Explain. Movement up a concentration gradient? Explain. __________________________________________________________________ __________________________________________________________________ __________________________________________________________________ __________________________________________________________________ __________________________________________________________________ __6. The movement of molecules is driven by the concentration gradient that exists between the two sides. A good definition of concentration gradient is "the change in concentration of a solute molecule over a distance between two points." One point is at the high end of the gradient, the other point is at the low end. In this module, we will be dealing primarily with very short distances, from one side of the cell membrane to the other, so the main focus will be upon concentration differences. __ 7. The direction and rate of net movement due to diffusion depends largely on the differences in concentration for a molecule on the two sides of the membrane. The larger the difference in concentration, the faster the rate of movement can be across the membrane. __ 8. In Figure 1.2, how many concentration gradients can you identify? ___________ . __ 9. If a cell membrane is permeable to solute particles, some will move across to the adjacent compartment. In the Figure 1.2, you can see some tiny solute molecules between the larger phospholipid molecules of the membrane, possibly on their ways from one side to the other. Add arrows to a few of the small and medium sized molecules to indicate in what direction you think they are heading. Explain your prediction. Explain your predictions in Fig 1.3 __________________________________________________________________ __________________________________________________________________ __________________________________________________________________ __________________________________________________________________ __________________________________________________________________ __ 10. Electrical gradients are especially important in moving charged particles into or out of a cell. You may remember from basic principles of magnetism that like charges repel one another, while opposite charges attract. In an electrical gradient, like that across the cell membrane, there are