The Molecular Biology of Plant Cells "d0e3313"

2.4.2—
Criteria For Active ('Uphill') Transport

To decide whether or not an ion or solute is actively transported across a membrane we need to know its activity or concentration in the two solutions separated by the membrane and for ions, in addition, we must know the electrical potential difference across the membrane. It is frequently difficult to measure the concentrations, and more difficult to measure the activity, of substances within cells with any accuracy, especially in those of higher plants.

In the giant cells of several sorts of algae, which may be 5,000 to 10,000 times the volume of a parenchyma cell in a root, such measurements are made

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Figure 2.12
An explanation of the way in which a decrease in electrical potential, y , across a membrane
can result in the diffusion of an ion against a gradient of concentration. Note that, in the initial
situation, in spite of concentration of j being greater in B, the electrochemical. potential gradient is
still directed 'downhill' towards B. As B fills with ion j, µ_j flattens out and, at equilibrium becomes zero—
at this point the 'uphill' concentration gradient and the 'downhill' electrical gradient are equal and opposite.

routinely. The electrical potential difference across membranes can be measured if a small glass micro-electrode, with a tip diameter of 1 to 3m m, can be inserted into the membrane-bounded compartment (see Clarkson, 1974).

Having made the necessary measurements a simple test can be applied to see if a given ion or solute within the compartment is at a higher or lower potential than in the surrounding solution. The principal snag in this analysis is that the cell or compartment should be in a steady state and that no net movement of solute should be occurring. In nature this condition is infrequently met.

Let us suppose that the ion j is at electrochemical equilibrium between the two compartments i.e.:

re-writing equation 2.3 and cancelling out

we have

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gathering the electrical terms to the left-hand side we get

yⁱⁿ —y^out is the electrical potential difference across the membrane where the ion j is at equilibrium and is given a special name, the Nernst Potential, and is usually symbolized Ej^N . We now compare this calculated equilibrium potential with the potential difference which is actually measured by the electrodes on either side of the membrane. If the calculated and observed values coincide we would conclude that, in spite of any differences in C_j across the membrane, the system was at equilibrium. If, however, the observed potential was lower than the equilibrium potential we would conclude that the electrical driving force was not sufficiently large to support the observed asymmetry of C_j and we would suspect that active transport was occurring. The example worked out in Table 2.8 may make this clearer. For each ion the appropriate Nernst Potential has

Table 2.8. Comparison of Nernst Potentials and equilibrium concentrations of ions with observed values from a hypothetical cell
Observed potential E = – 116 mV Temperature 293°K (20°C)
	Concentration		E_jⁿ (mV)	E_j (mV)	Equilibrium concentrat (mM )
Ion	Outside (mM )	Inside	E_jⁿ (mV)	E_j (mV)	Equilibrium concentrat (mM )
K⁺	1	100	–116	0	100
Na⁺	10	50	–41	–75	1,000
Cl^–	11	150	+65	–181	0.11
The Nernst Potential for each ion was calculated from equation 2.5. The column E_j is the difference between the Nernst Potential and the observed potential. Where the observed potential E is the same as the Nernst Potential the ion is at electrochemical equilibrium. If the sign of this difference is negative it means that, for cations, the electrochemical potential is 'downhill' into the cell; for anions, however, it signifies an 'uphill' gradient. In the last column, the equilibrium concentration inside the cell, has been computed on the assumption that the observed potential was equal to the Nernst Potential for each ion. Departures between this computed value and the observed concentrations indicate how far the cell is from being in a passive equilibrium with respect to the ion.

been calculated from the observed concentrations on the outside and inside of the membrane using equation 2.5. The exact correspondence of the Nernst Potential for potassium,

, with the measured electric potential difference shows that potassium ions inside the cell are at electrochemical equilibrium with those outside. For sodium, however,

is much less negative than E, hence Na⁺ is at a lower electrochemical potential inside; there is, therefore, a strong tendency for sodium to diffuse into the cell, and the fact that the observed

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concentration is only 1/20 th of the equilibrium concentration strongly suggests that metabolic energy must be coupled to a Na⁺ -efflux pump. Chloride ions in the cell are a very long way indeed from being in electrochemical equilibrium with their surroundings, being more than 1,000 times greater than the equilibrium concentration, thus their movement into the cell is steeply uphill.

In theory this type of analysis can be applied to any ion, although it is difficult to apply to minor ionic constituents, e.g. trace elements, because they may be complexed with organic ligands within the cell so that their ionic activity may be very much lower than their concentration as measured by chemical analysis.

If an analysis of the kind described in Table 2.8 shows that the transport of an ion in a given direction is 'uphill', one should not conclude necessarily that the membrane is equipped with a special pumping mechanism for that ion. In some cases it may be, but in others the 'uphill' transport of the ion may be coupled with the 'downhill' transport of another via a common carrier; this latter possibility is described under the heading Co-transport on p.56.

2.4.2— Criteria For Active ('Uphill') Transport

2.4.2—
Criteria For Active ('Uphill') Transport