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Plasmalemmal H+ extruders in mammalian alveolar macrophages
Thomas A Heming1, Akhil Bidani
1Pulmonary and Critical Care Medicine, Department of Internal Medicine, University of Texas Health Science Center, Houston, TX 77225-0708, USA. Thomas.Heming@uth.tmc.edu
Abstract:
The distribution of plasmalemmal V-type H+-pumps (V-ATPase) among mammalian macrophages (mvarphi) is uncertain and, hence, the functional significance of mvarphi plasmalemmal V-ATPase is unclear. This study investigated the role of V-ATPase in the regulation of intracellular pH (pH(i)) by resident alveolar mvarphi from sheep, pigs, dogs and rabbits. The fluorescent probe 2',7'-biscarboxyethyl-5,6-carboxyfluorescein was used to monitor baseline pH(i) and the rate of pH(i) recovery (dpH(i)/dt) from intracellular acid-loads (NH(4)Cl prepulse). Baseline pH(i) was 7.1-7.2. In sheep, pig and dog studies, 10 microM bafilomycin A(1) (a selective V-ATPase inhibitor) caused a rapid fall in baseline pH(i) (0.15-0.20 units); baseline values were unaffected by 0.1 mM amiloride (a Na+ transport inhibitor). V-ATPase activity (bafilomycin-sensitive component of dpH(i)/dt) was solely responsible for pH(i) recovery from intracellular acid-loads at acid-loaded pH(i) values >6.8-6.9. Na+/H+ exchange (amiloride-sensitive component of dpH(i)/dt) was detected only at acid-loaded pH(i) values <6.8. The activity of both H+ extruders increased at lower pH(i) values, albeit the Na+/H+ exchanger was more pH-sensitive than was V-ATPase. In rabbit studies, 10 microM bafilomycin A(1) and 1 mM N-ethylmaleimide (a non-specific H+-pump inhibitor) produced similar falls in baseline mvarphi pH(i), but had significantly larger effects than did the selective V-ATPase inhibitor concanamycin A (
Insights
Plasmalemmal V-type H+-pumps (V-ATPase) are crucial for regulating intracellular pH in alveolar macrophages. These pumps, along with Na+/H+ exchange, maintain pH balance in various mammals.
Area of Science:
- Cell Biology
- Physiology
- Biochemistry
Background:
- The role of V-type H+-pumps (V-ATPase) in the plasma membrane of mammalian macrophages (mφ) and their functional significance remain poorly understood.
- Intracellular pH (pH(i)) regulation is vital for macrophage function.
Purpose of the Study:
- To investigate the role of V-ATPase in regulating intracellular pH (pH(i)) in resident alveolar macrophages from sheep, pigs, dogs, and rabbits.
- To compare the contribution of V-ATPase and Na+/H+ exchange to pH(i) homeostasis.
Main Methods:
- Utilized the fluorescent probe 2',7'-biscarboxyethyl-5,6-carboxyfluorescein to monitor baseline pH(i) and pH(i) recovery rates.
- Employed selective V-ATPase inhibitors (bafilomycin A1, concanamycin A) and a Na+ transport inhibitor (amiloride) to assess transporter activity.
- Induced intracellular acid-loads using an NH4Cl prepulse to study pH(i) recovery mechanisms.
Main Results:
- Baseline pH(i) in alveolar macrophages was consistently around 7.1-7.2 across species.
- V-ATPase activity was the primary mechanism for pH(i) recovery from acid-loads at pH(i) >6.8-6.9 in sheep, pig, and dog macrophages.
- Na+/H+ exchange contributed to pH(i) recovery only at more acidic pH(i) values (<6.8) and exhibited higher pH sensitivity than V-ATPase.
- In rabbits, V-ATPase and non-specific H+-pump inhibitors showed similar effects on baseline pH(i), suggesting a significant V-ATPase role.
Conclusions:
- Plasmalemmal V-ATPase plays a major role in the intracellular pH regulation of alveolar macrophages in sheep, pigs, dogs, and rabbits.
- The relative contributions of V-ATPase and Na+/H+ exchange to pH(i) homeostasis vary with the degree of intracellular acidification.
- Understanding these mechanisms is crucial for comprehending macrophage physiology and function.