Medical Physiology, 3rd Edition

pH Regulation of Intracellular Fluid

Clinicians focus on the acid-base status of blood plasma, the fluid compartment whose acid-base status is easiest to assess. However, the number of biochemical reactions and other processes that occur outside a cell pales in comparison to the number of those that occur inside. It stands to reason that the most important biological fluid for pH regulation is the cytosol. It is possible to monitor intracellular pH (pHi) continuously in isolated cells and tissues using pH-sensitive dyes or microelectrodes. Moreover, using magnetic resonance techniques, one can monitor pHi in living humans (e.g., in skeletal muscle). The pH of cells both influences, and is influenced by, extracellular pH.

Ion transporters at the plasma membrane closely regulate the pH inside of cells

Figure 28-12A shows a hypothetical cell with three acid-base transporters: an electroneutral Na/HCO3 cotransporter (NBCn1; see p. 122) a Na-H exchanger (NHE1; pp. 124–125), and a Cl-HCO3 or anion exchanger (AE2; see p. 124). We will use NBCn1 and NHE1 as prototypic acid extruders, transporters that tend to raise intracellular pH (pHi). Both use the energy of the Na+ gradient to either import image or export H+ from the cytosol. We will take the Cl-HCO3 exchanger AE2 as the prototypic acid loader, a transporter that tends to lower pHi. AE2, driven by the steep out-to-in Cl gradient, moves image out of the cell. The intracellular acid loading produced by Cl-HCO3 exchange is a chronic acid load because it tends to acidify the cell as long as the transporter is active. In our example, the chronic acid extrusion via NBCn1 and NHE1 balances the chronic acid loading via AE2, producing a steady state. Several other acid extruders and acid loaders may contribute to pHi regulation (see p. 127), and each cell type has a characteristic complement of such transporters.

image

FIGURE 28-12 Recovery of a cell from intracellular acid and alkali loads. CA II, carbonic anhydrase II.

Imagine that we use a micropipette to inject HCl into the cell in Figure 28-12A. This injection of an acute acid load—a one-time-only introduction of a fixed quantity of acid—causes an immediate fall in pHi(see Fig. 28-12B, black curve), an example of intracellular metabolic acidosis. If we add HCl to a beaker, the pH remains low indefinitely. However, when we acutely acid-load a cell, pHi spontaneously recovers to the initial value. This pHi recovery cannot be due to passive H+ efflux out of the cell, inasmuch as the electrochemical gradient for H+ (see p. 127) usually favors H+ influx. Hence, the pHi recovery reflects the active transport of acid from the cell—a metabolic compensation to a metabolic acid load.imageN28-10

N28-10

Acid Extruders and Acid Loaders in a Cell

Contributed by Emile Boulpaep, Walter Boron

Of the acid extruders that can produce pHi recovery, the most widely distributed is the NHE1 isoform of the Na-H exchanger. However, mechanisms that mediate image uptake often coexist with the Na-H exchanger in the same cell and are sometimes far more powerful than the Na-H exchanger. These image transporters include the Na+-driven Cl-HCO3 exchanger (or NDCBE, illustrated in Fig. 5-14, no. 17) and the Na/HCO3 cotransporters with Na+:image stoichiometries of 1 : 2 (NBCe1 and NBCe2, the electrogenic Na/HCO3 cotransporters) and 1 : 1 (NBCn1, the electroneutral Na/HCO3 cotransporter). Note that the electrogenic Na/HCO3 cotransporter that operates with an Na+:image stoichiometry of 1 : 3 (see Fig. 5-14, no. 19) mediates net image efflux—which reflects the ionic and electrical gradients that govern its thermodynamic properties—and functions as an acid loader. However, the Na/HCO3 cotransporters with Na+:image stoichiometries of 1 : 2 and 1 : 1 both mediate the net uptake of image and function as acid extruders.

Other transporters that extrude acid from cells include the V-type H pump and the H-K exchange pump.

Other transport processes that acid-load cells include the passive influx of H+ as well as the efflux of image either through channels (e.g., gamma-aminobutyric acid or glycine-activated Cl channels) or through the K/HCO3 cotransporter.

For a more detailed discussion of these transporters, see page 127.

During the recovery of pHi from the acid load in Figure 28-12B, NBCn1 and NHE1not only must extrude the quantity of H+ previously injected into the cell (the acute acid load), they also must counteract whatever acid loading AE2 continues to impose on the cell (the chronic acid load).

Acid-extrusion rates tend to be greatest at low pHi values and fall off as pHi rises (see Fig. 28-12C, black curve). As discussed below, low pHi also inhibits AE2 and thus acid loading. The response to low pHitherefore involves two feedback loops operating in push-pull fashion, stimulating acid extrusion and inhibiting acid loading.

In addition to pHi, hormones, growth factors, oncogenes, cell volume, and extracellular pH (pHo) all can modulate these transporters. The pHo effect is especially important. In general, a low pHo slows the rate of pHi recovery from acute acid loads and reduces the final steady-state pHi (see Fig. 28-12B, red curve), whereas a high pHo does the opposite (blue curve). The underlying cause of these pHo effects is a shift of the pHi dependence of acid extrusion (see Fig. 28-12C, red and blue curves) and, as we shall see, changes in acid loading.

Cells also spontaneously recover from acute alkaline loads. If we inject a cell with potassium hydroxide (KOH) (see Fig. 28-12D), pHi rapidly increases but then slowly recovers to its initial value (see Fig. 28-12E, black curve), which reflects stimulation of acid loading and inhibition of acid extrusion. The injection of OH represents a metabolic alkalosis, whereas increased Cl-HCO3 exchange represents a metabolic compensation. imageN28-10 The Cl-HCO3 exchanger is most active at high pHi values (see Fig. 28-12F). As discussed previously, increases in pHi generally inhibit acid extruders. Thus, the response to alkaline loads—like the response to acid loads—includes dual feedback loops operating in push-pull fashion, stimulating acid loading and inhibiting acid extrusion. During the pHi recovery from an acute alkaline load, Cl-HCO3 exchange not only must neutralize the alkali previously injected into the cell (the acute alkali load), but also must counteract the alkalinizing effect of acid extruders such as the Na/HCO3 cotransporter.

Like acid extruders, Cl-HCO3 exchangers are under the control of hormones and growth factors with a pHo sensitivity that is opposite to that of the acid extruders (i.e., low pHo enhances Cl-HCO3 exchange). In general, an extracellular metabolic acidosis, with its simultaneous fall in both pHo and [image] (see Table 28-3), stimulates acid loading via Cl-HCO3 exchange (see Fig. 28-12F, red curve). During the recovery from acute alkali loads, metabolic acidosis therefore causes pHi to fall more rapidly and to reach a lower steady-state value (see Fig. 28-12E, red curve).

Indirect interactions between K+ and H+ make it appear as if cells have a K-H exchanger

Clinicians have long known that extracellular acidosis leads to a release of K+ from cells, and thus hyperkalemia (see p. 795). Conversely, hyperkalemia leads to the exit of H+ from cells, resulting in extracellular acidosis. These observations, which one can duplicate in single, isolated cells, led to the suggestion that cells generally have K-H exchangers. It is true that specialized cells in the stomach (see pp. 865–866) and kidney (see pp. 827–828) do have an ATP-driven pump that extrudes H+ in exchange for K+. Moreover, the K/HCO3 cotransporter described in certain cells could mimic a K-H exchanger. Nevertheless, the effects that long ago led to the K-H–exchange hypothesis probably reflect indirect interactions of H+ and K+.

One example of apparent K-H exchange is that hyperkalemia causes an intracellular alkalosis. Not only is this effect not due to a 1 : 1 exchange of K+ for H+, it is not even due to the increase in [K+]o per se. Instead, the high [K+]o depolarizes the cell, and this positive shift in membrane voltage can promote events such as the net uptake of image via the electrogenic Na/HCO3 cotransporter (see p. 122) and thus a rise in pHi—a depolarization-induced alkalinization. Depolarization also can indirectly stimulate other transporters that alkalinize the cell. We discuss the converse example of acidemia causing hyperkalemia on page 795: Extracellular acidosis lowers pHi and inhibits transporters responsible for K+ uptake, which leads to net K+ release from cells. Although we have no evidence that a K-H exchanger exists in humans, imagining that it exists is sometimes a helpful tool for quickly predicting interactions of H+ and K+ in a clinical setting.

Changes in intracellular pH are often a sign of changes in extracellular pH, and vice versa

By definition, acid extrusion (e.g., Na/HCO3 cotransport) equals acid loading (e.g., Cl-HCO3 exchange) in the steady state. Disturbing this balance shifts pHi. For example, extracellular metabolic acidosis inhibits acid extrusion and stimulates acid loading, thus lowering steady-state pHi. This intracellular acidosis is not instantaneous, but develops over a minute or two. Conversely, extracellular metabolic alkalosis leads to a slow increase in steady-state pHi. Generally, a change in pHo shifts pHi in the same direction, but ΔpHi is usually only 20% to 60% of ΔpHo. In other words, an extracellular metabolic acidosis causes a net transfer of acid from the extracellular to the intracellular space, and an extracellular metabolic alkalosis has the opposite effect. Thus, cells participate in buffering extracellular acid and alkali loads.

Extracellular respiratory acidosis generally affects pHi in three phases. First, the increase in extracellular [CO2]Dis creates an inwardly directed gradient for CO2. This dissolved gas rapidly enters the cell (Fig. 28-13A) and produces image and H+. This intracellular respiratory acidosis manifests itself as a rapid fall in pHi (see Fig. 28-13B, phase A in lower panel). Carbonic anhydrase imageN18-3 greatly accelerates the formation of image and H+ from CO2, so that pHi can complete its decline in just a few seconds.

image

FIGURE 28-13 Response of cell to extracellular respiratory acidosis.

The cell recovers from this acid load, but only feebly (see Fig. 28-13B, phase B in lower panel) because the decrease in pHo inhibits acid extrusion and stimulates acid loading. Over a period of minutes, pHi may recover only partially if at all.

Finally, the extracellular respiratory acidosis stimulates the kidneys to upregulate acid-base transporters and thus stimulate urinary acid secretion (see p. 832). The net effect, over a period of hours or days, is an extracellular metabolic compensation that causes pHo to increase gradually (see Fig. 28-13B, phase C in upper panel). As pHo rises, it gradually relieves the inhibition of NBCn1 and NHE1 and other acid extruders and cancels the stimulation of Cl-HCO3 exchange and other acid loaders. Thus, intracellular pH recovers in parallel with extracellular pH, albeit by only 20% to 60% as much as pHo (see Fig. 28-13B, phase C in lower panel). The pHi recovery represents an intracellular metabolic compensation to the intracellular respiratory acidosis.

Not all cell types have the same resting pHi. Moreover, different cell types often have very different complements of acid-base transporters and very different ways of regulating these transporters. Nevertheless, the example in Figure 28-13 illustrates that the fate of pHi is closely intertwined with that of interstitial fluid, and thus blood plasma. During respiratory acid-base disturbances, the lungs generate the insult, and virtually every other cell in the body must defend itself against it. In the case of metabolic acid-base disturbances, however, some cells may generate the insult, whereas the others attempt to defend themselves against it. From a teleological perspective, one can imagine that the primary reason that the body regulates the pH of the blood plasma and extracellular fluids is to allow the cells to properly regulate their pHi. The primary reason why the clinical assessment of blood acid-base parameters can be useful is that these parameters tend to parallel cellular acid-base status.