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Extracellular fluid flow and chloride content modulate H(+) transport by osteoclasts.

Morethson P - BMC Cell Biol. (2015)

Bottom Line: Bone resorption takes place within the basic multicellular units (BMU), and the surface to be resorbed is isolated from adjacent bone surfaces by a sealing zone between osteoclast membrane and bone matrix, which defines the limits of the resorption lacuna.Considering that the extracellular fluid (ECF) in both BMU and the resorption lacuna can be isolated from its surroundings, I hypothesize that flow and ion composition of the bone ECF in these sites might contribute to the regulation of osteoclast H(+) secretion.The data suggest, for the first time, that ECF flow and Cl(-) content have direct effects on osteoclast H(+) secretion and could be part of a mechanism determining the onset of osteoclast H(+) secretion required for bone resorption.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology and Biophysics, Institute of Biomedical Sciences, University of São Paulo, São Paulo, SP, Brazil. primoret@icb.usp.br.

ABSTRACT

Background: Bone resorption takes place within the basic multicellular units (BMU), and the surface to be resorbed is isolated from adjacent bone surfaces by a sealing zone between osteoclast membrane and bone matrix, which defines the limits of the resorption lacuna. Considering that the extracellular fluid (ECF) in both BMU and the resorption lacuna can be isolated from its surroundings, I hypothesize that flow and ion composition of the bone ECF in these sites might contribute to the regulation of osteoclast H(+) secretion. To investigate this hypothesis, I evaluated the H(+) secretion properties of individual osteoclasts and osteoclast-like cells (OCL-cells) and investigated whether changes in flow or chloride content of the extracellular solution modify the H(+) secretion properties in vitro.

Results: The results show that 1) osteoclasts are unable to secrete H(+) and regulate intracellular pH (pHi) under continuous flow conditions and exhibit progressive intracellular acidification; 2) the cessation of flow coincides with the onset of H(+) secretion and subsequent progressive intracellular alkalinization of osteoclasts and OCL-cells; 3) osteoclasts exhibit spontaneous rhythmic oscillations of pHi in non-flowing ECF, 4) pHi oscillations are not abolished by concanamycin, NPPB, or removal of extracellular Na(+) or Cl(-); 5) extracellular Cl(-) removal modifies the pattern of oscillations, by diminishing H(+) secretion; 6) pHi oscillations are abolished by continuous flowing of ECF over osteoclasts and OCL-cells.

Conclusions: The data suggest, for the first time, that ECF flow and Cl(-) content have direct effects on osteoclast H(+) secretion and could be part of a mechanism determining the onset of osteoclast H(+) secretion required for bone resorption.

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Effect of inhibitors of H+-secreting proteins in the oscillating intracellular pH (pHi) of primary osteoclasts under non-flowing standard HEPES-buffered solution. a. The pHi oscillations were not abolished by applying a zero Na+ solution (0 Na+), inhibitor of Na+/H+ exchanger. b. The pHi oscillations were not abolished in the presence of concanamycin (Conc.), inhibitor of H+-ATPase. (Nig. = nigericin clamps pHi at 7.0.). c. The pHi oscillations were not abolished in the presence of NPPB, inhibitor of Clˉ channels. d. The pHi oscillations were not abolished by applying a zero Clˉ solution (0 Clˉ), inhibitor of Clˉ transporting proteins; however there is a noticeable and progressive intracellular acidification from one cycle to the next following the removal of extracellular Clˉ (0 Clˉ). e. Parameters applied for the analyses of the oscillating pHi
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Fig4: Effect of inhibitors of H+-secreting proteins in the oscillating intracellular pH (pHi) of primary osteoclasts under non-flowing standard HEPES-buffered solution. a. The pHi oscillations were not abolished by applying a zero Na+ solution (0 Na+), inhibitor of Na+/H+ exchanger. b. The pHi oscillations were not abolished in the presence of concanamycin (Conc.), inhibitor of H+-ATPase. (Nig. = nigericin clamps pHi at 7.0.). c. The pHi oscillations were not abolished in the presence of NPPB, inhibitor of Clˉ channels. d. The pHi oscillations were not abolished by applying a zero Clˉ solution (0 Clˉ), inhibitor of Clˉ transporting proteins; however there is a noticeable and progressive intracellular acidification from one cycle to the next following the removal of extracellular Clˉ (0 Clˉ). e. Parameters applied for the analyses of the oscillating pHi

Mentions: The inhibition of the Na+/H+ exchanger by applying ECF containing zero sodium (0 Na+) (n = 5), the inhibition of H+-ATPase by concanamycin (n = 3) (Fig. 4a and b) or of H+ channels by Zn2+ (n = 2) did not disrupt or modify the oscillatory pattern of pHi in osteoclasts. Thus, these H+-transporting proteins do not appear to participate in pH regulation by osteoclasts and OCL-cells.Fig. 4


Extracellular fluid flow and chloride content modulate H(+) transport by osteoclasts.

Morethson P - BMC Cell Biol. (2015)

Effect of inhibitors of H+-secreting proteins in the oscillating intracellular pH (pHi) of primary osteoclasts under non-flowing standard HEPES-buffered solution. a. The pHi oscillations were not abolished by applying a zero Na+ solution (0 Na+), inhibitor of Na+/H+ exchanger. b. The pHi oscillations were not abolished in the presence of concanamycin (Conc.), inhibitor of H+-ATPase. (Nig. = nigericin clamps pHi at 7.0.). c. The pHi oscillations were not abolished in the presence of NPPB, inhibitor of Clˉ channels. d. The pHi oscillations were not abolished by applying a zero Clˉ solution (0 Clˉ), inhibitor of Clˉ transporting proteins; however there is a noticeable and progressive intracellular acidification from one cycle to the next following the removal of extracellular Clˉ (0 Clˉ). e. Parameters applied for the analyses of the oscillating pHi
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4536797&req=5

Fig4: Effect of inhibitors of H+-secreting proteins in the oscillating intracellular pH (pHi) of primary osteoclasts under non-flowing standard HEPES-buffered solution. a. The pHi oscillations were not abolished by applying a zero Na+ solution (0 Na+), inhibitor of Na+/H+ exchanger. b. The pHi oscillations were not abolished in the presence of concanamycin (Conc.), inhibitor of H+-ATPase. (Nig. = nigericin clamps pHi at 7.0.). c. The pHi oscillations were not abolished in the presence of NPPB, inhibitor of Clˉ channels. d. The pHi oscillations were not abolished by applying a zero Clˉ solution (0 Clˉ), inhibitor of Clˉ transporting proteins; however there is a noticeable and progressive intracellular acidification from one cycle to the next following the removal of extracellular Clˉ (0 Clˉ). e. Parameters applied for the analyses of the oscillating pHi
Mentions: The inhibition of the Na+/H+ exchanger by applying ECF containing zero sodium (0 Na+) (n = 5), the inhibition of H+-ATPase by concanamycin (n = 3) (Fig. 4a and b) or of H+ channels by Zn2+ (n = 2) did not disrupt or modify the oscillatory pattern of pHi in osteoclasts. Thus, these H+-transporting proteins do not appear to participate in pH regulation by osteoclasts and OCL-cells.Fig. 4

Bottom Line: Bone resorption takes place within the basic multicellular units (BMU), and the surface to be resorbed is isolated from adjacent bone surfaces by a sealing zone between osteoclast membrane and bone matrix, which defines the limits of the resorption lacuna.Considering that the extracellular fluid (ECF) in both BMU and the resorption lacuna can be isolated from its surroundings, I hypothesize that flow and ion composition of the bone ECF in these sites might contribute to the regulation of osteoclast H(+) secretion.The data suggest, for the first time, that ECF flow and Cl(-) content have direct effects on osteoclast H(+) secretion and could be part of a mechanism determining the onset of osteoclast H(+) secretion required for bone resorption.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology and Biophysics, Institute of Biomedical Sciences, University of São Paulo, São Paulo, SP, Brazil. primoret@icb.usp.br.

ABSTRACT

Background: Bone resorption takes place within the basic multicellular units (BMU), and the surface to be resorbed is isolated from adjacent bone surfaces by a sealing zone between osteoclast membrane and bone matrix, which defines the limits of the resorption lacuna. Considering that the extracellular fluid (ECF) in both BMU and the resorption lacuna can be isolated from its surroundings, I hypothesize that flow and ion composition of the bone ECF in these sites might contribute to the regulation of osteoclast H(+) secretion. To investigate this hypothesis, I evaluated the H(+) secretion properties of individual osteoclasts and osteoclast-like cells (OCL-cells) and investigated whether changes in flow or chloride content of the extracellular solution modify the H(+) secretion properties in vitro.

Results: The results show that 1) osteoclasts are unable to secrete H(+) and regulate intracellular pH (pHi) under continuous flow conditions and exhibit progressive intracellular acidification; 2) the cessation of flow coincides with the onset of H(+) secretion and subsequent progressive intracellular alkalinization of osteoclasts and OCL-cells; 3) osteoclasts exhibit spontaneous rhythmic oscillations of pHi in non-flowing ECF, 4) pHi oscillations are not abolished by concanamycin, NPPB, or removal of extracellular Na(+) or Cl(-); 5) extracellular Cl(-) removal modifies the pattern of oscillations, by diminishing H(+) secretion; 6) pHi oscillations are abolished by continuous flowing of ECF over osteoclasts and OCL-cells.

Conclusions: The data suggest, for the first time, that ECF flow and Cl(-) content have direct effects on osteoclast H(+) secretion and could be part of a mechanism determining the onset of osteoclast H(+) secretion required for bone resorption.

Show MeSH