INVESTIGADORES
LAROCCA Maria Cecilia
artículos
Título:
Hepatocyte Membrane Water Permeability Measured
Autor/es:
GRADILONE, S.A.; OCHOA, E.J.; GARCIA, F; LAROCCA, MC; PELLEGRINO, J.M.; MARINELLI, R.A.
Revista:
ANALYTICAL BIOCHEMISTRY
Editorial:
Elsevier
Referencias:
Año: 2002 vol. 302 p. 104 - 107
ISSN:
0003-2697
Resumen:
We previously found that hepatocytes are able to
control their osmotic membrane water permeability
(Pf) by regulating the number of surface aquaporin
water channels. Hepatocyte Pf has been assessed by
phase-contrast microscopy and cell image analysis, an
established but relatively laborious procedure. We report
here an alternative method to assess hepatocyte
Pf) by regulating the number of surface aquaporin
water channels. Hepatocyte Pf has been assessed by
phase-contrast microscopy and cell image analysis, an
established but relatively laborious procedure. We report
here an alternative method to assess hepatocyte
Pf has been assessed by
phase-contrast microscopy and cell image analysis, an
established but relatively laborious procedure. We report
here an alternative method to assess hepatocyte
Pf based on a single silicone layer filtering centrifugation
system. Isolated rat hepatocytes were incubated
in hypotonic or isotonic buffers containing 3H2O as a
tracer and, then, were filtered by rapid centrifugation
through a silicone layer down to a lysis layer. Osmotically
driven radioactivity (i.e., 3H2O) within hepatocytes
was calculated as the difference between the
dpm in lysis media measured under hypotonic and
isotonic conditions. The Pf calculated from the initial
slope of the radioactivity-versus-time curve was 18
f based on a single silicone layer filtering centrifugation
system. Isolated rat hepatocytes were incubated
in hypotonic or isotonic buffers containing 3H2O as a
tracer and, then, were filtered by rapid centrifugation
through a silicone layer down to a lysis layer. Osmotically
driven radioactivity (i.e., 3H2O) within hepatocytes
was calculated as the difference between the
dpm in lysis media measured under hypotonic and
isotonic conditions. The Pf calculated from the initial
slope of the radioactivity-versus-time curve was 18
3H2O as a
tracer and, then, were filtered by rapid centrifugation
through a silicone layer down to a lysis layer. Osmotically
driven radioactivity (i.e., 3H2O) within hepatocytes
was calculated as the difference between the
dpm in lysis media measured under hypotonic and
isotonic conditions. The Pf calculated from the initial
slope of the radioactivity-versus-time curve was 18
3H2O) within hepatocytes
was calculated as the difference between the
dpm in lysis media measured under hypotonic and
isotonic conditions. The Pf calculated from the initial
slope of the radioactivity-versus-time curve was 18
Pf calculated from the initial
slope of the radioactivity-versus-time curve was 18
mm/s at 4°C. Hepatocytes treated with dibutyryl cyclic
AMP, to increase Pf through the plasma membrane
insertion of aquaporins, showed an increased Pf value
of 37 mm/s. The aquaporin blocker dimethyl sulfoxide
selectively prevented the agonist-induced hepatocyte
Pf. These data are in good agreement with the corresponding
values determined by quantitative phasecontrast
microscopy; thus, the method developed allows
the rapid and reliable measurement of hepatocyte
f. These data are in good agreement with the corresponding
values determined by quantitative phasecontrast
microscopy; thus, the method developed allows
the rapid and reliable measurement of hepatocyte
Pf.f.
Pf through the plasma membrane
insertion of aquaporins, showed an increased Pf value
of 37 mm/s. The aquaporin blocker dimethyl sulfoxide
selectively prevented the agonist-induced hepatocyte
Pf. These data are in good agreement with the corresponding
values determined by quantitative phasecontrast
microscopy; thus, the method developed allows
the rapid and reliable measurement of hepatocyte
f. These data are in good agreement with the corresponding
values determined by quantitative phasecontrast
microscopy; thus, the method developed allows
the rapid and reliable measurement of hepatocyte
Pf.f.
Pf value
of 37 mm/s. The aquaporin blocker dimethyl sulfoxide
selectively prevented the agonist-induced hepatocyte
Pf. These data are in good agreement with the corresponding
values determined by quantitative phasecontrast
microscopy; thus, the method developed allows
the rapid and reliable measurement of hepatocyte
f. These data are in good agreement with the corresponding
values determined by quantitative phasecontrast
microscopy; thus, the method developed allows
the rapid and reliable measurement of hepatocyte
Pf.f.
m/s at 4°C. Hepatocytes treated with dibutyryl cyclic
AMP, to increase Pf through the plasma membrane
insertion of aquaporins, showed an increased Pf value
of 37 mm/s. The aquaporin blocker dimethyl sulfoxide
selectively prevented the agonist-induced hepatocyte
Pf. These data are in good agreement with the corresponding
values determined by quantitative phasecontrast
microscopy; thus, the method developed allows
the rapid and reliable measurement of hepatocyte
f. These data are in good agreement with the corresponding
values determined by quantitative phasecontrast
microscopy; thus, the method developed allows
the rapid and reliable measurement of hepatocyte
Pf.f.
Pf through the plasma membrane
insertion of aquaporins, showed an increased Pf value
of 37 mm/s. The aquaporin blocker dimethyl sulfoxide
selectively prevented the agonist-induced hepatocyte
Pf. These data are in good agreement with the corresponding
values determined by quantitative phasecontrast
microscopy; thus, the method developed allows
the rapid and reliable measurement of hepatocyte
f. These data are in good agreement with the corresponding
values determined by quantitative phasecontrast
microscopy; thus, the method developed allows
the rapid and reliable measurement of hepatocyte
Pf.f.
Pf value
of 37 mm/s. The aquaporin blocker dimethyl sulfoxide
selectively prevented the agonist-induced hepatocyte
Pf. These data are in good agreement with the corresponding
values determined by quantitative phasecontrast
microscopy; thus, the method developed allows
the rapid and reliable measurement of hepatocyte
f. These data are in good agreement with the corresponding
values determined by quantitative phasecontrast
microscopy; thus, the method developed allows
the rapid and reliable measurement of hepatocyte
Pf.f.
mm/s. The aquaporin blocker dimethyl sulfoxide
selectively prevented the agonist-induced hepatocyte
Pf. These data are in good agreement with the corresponding
values determined by quantitative phasecontrast
microscopy; thus, the method developed allows
the rapid and reliable measurement of hepatocyte
f. These data are in good agreement with the corresponding
values determined by quantitative phasecontrast
microscopy; thus, the method developed allows
the rapid and reliable measurement of hepatocyte
Pf.f.