Cadmium transport in isolated zinc-cadmium competition BEVERLY Department and Public
perfused
S. KINGSLEY AND JOHN M. FRAZIER of Environmental Health Sciences, Johns Health, Baltimore, Maryland 21205
KINGSLEY, BEVERLY S., AND JOHN M. FRAZIER. Cadmium transport in isolated perfused rat Liver: zinc-cadmium competition. Am. J. Physiol, 236(3): C139-C143, 1979 or Am. J. Physiol.: Cell Physiol. 5(2): C139-C143, 1979.-The hypothesis that one component of cadmium uptake by rat hepatocytes involves a mediated transport pathway normally operative for zinc transport was tested in the isolated perfused rat liver preparation. Excess zinc in the perfusion medium suppressed cadmium uptake as indicated by the decrease in the normalized clearance (initial clearance divided by liver weight) from 0.340 t 0.019 (ml/min)/g in the presence of normal zinc concentrations (Zn:Cd molar ratio, 1.6) to 0.138 t 0.017 (ml/min)/g (Zn: Cd molar ratio, 13.0). In excess-zinc control experiments (no cadmium present) little zinc is accumulated by the liver, apparently due to competition between intrahepatic and extracellular binding. Exposure to cadmium increases both zinc secretion into the perfusion medium and biliary excretion of zinc. The effect at the sinusoidal membrane is probably a result of both the blockage of zinc resorption during cadmium uptake and the displacement of intrahepatic zinc. The effect on biliary excretion of zinc is due solely to displacement of intrahepatic zinc. These results are consistent with the proposed hypothesis for cadmium transport.
Hopkins
MATERIALS
AND
rat liver:
School
of Hygiene
METHODS
The recirculating isolated perfused rat liver preparation of Miller (6) was employed for these experiments. Male
Wistar
rats of 340 g average
weight
were
used
as
liver donors. Details of the surgical procedure were previously reported (4). The perfusion medium consisted of a Krebs-Ringer bicarbonate buffer supplemented with dextrose (0.8 g/l) and bovine serum albumin (4% wt/vol; fraction V; Sigma Chemical Corp., St. Louis, MO). Bovine serum albumin was carefully selected because the commercial product is often contaminated with zinc, copper, and iron. All experiments were performed with an initial zinc concentration of 0,66-0.90 lug Zn/ml, which approximates the normal labile serum zinc concentration in these rats determined in this laboratory. Initial volume of the perfusion medium was 350 ml and perfusion pressure was maintained at 17.5 cmHz0, corresponding to a perfusion rate of 60-75 ml/min in control experiments. The pH of the perfusion medium was maintained between 7.35-7.45 by additions of sodium bicarbonate. Each 5-h experiment was performed according to the following protocol. The first 60 min served as a control biliary excretion; mediated transport period. At 60 min (t = 0) four different treatments were imposed (four liver preparations per treatment group): 1) normal zinc control (no additions), 2) excess zinc control HEPATXC TRANSPORT OF CADMIUM is controlled by the (addition of zinc, as ZnSOd, to perfusion medium, resulting in a total zinc concentration of 8.4 lug Zn/ml or 129 following processes: 1) the transport of cadmium across FM), 3) normal zinc plus cadmium (addition of cadmium, the sinusoidal membrane of the hepatocytes, 2) intracelas CdClz, giving a concentration of 1.0 lug Cd/ml or 8.94 lular binding to cell organelles and cytoplasmic macromolecules, and 3) transport across the membrane sepa- PM and a Zn:Cd molar ratio of 1,6), and 4) excess zinc (addition of both zinc and cadmium rerating the hepatocyte from the biliary caraliculi. These plus cadmium sulting in Zn:Cd molar ratio of 13.0). At fixed intervals processes are of particular importance because the liver periods, samplays a central role in the distribution and excretion of during both the control and experimental ples of perfusion medium were collected for cadmium cadmium (5). Previous studies in this laboratory (4) have characterized cadmium-uptake kinetics in the isolated and zinc analysis. Cumulative samples of bile over l-h perfused rat liver. These studies suggested that both intervals were also collected throughout the experiment simple diffusion and mediated transport are involved in and analyzed for cadmium and zinc. Metal analyses of perfusion medium, bile, and liver cadmium transport into the hepatocytes at the sinusoidal tissue were performed by atomic absorption spectrophomembrane and that the mediated transport component may involve a pathway normally utilized for zinc trans- tometry (AA-5, Varian Techtron, Victoria, Australia) as port. The present investigation was designed to test this previously described (4). Data are reported as X t SE, rz = 4, unless otherwise indicated. Comparisons for statishypothesis in the isolated perfused rat liver preparation. If cadmium and zinc use the same membrane carrier for tical significance were performed by Student’s t test with a level of significance of P 5 0,05. uptake, then excess extracellular zinc should inhibit cadmium uptake by competing for the carrier on the sinusRESULTS oidal side of the hepatocyte membrane. Observations of the kinetics of cadmium and zinc in the perfusion medium Kinetics of cadmium and zinc in perfusion medium. and the biliary excretion of these two metals are reported. The clearance of cadmium from the perfusion medium 0363-6143/79/0000-OUOO$O1.25
Copyright 0 1979 the American Physiological Society Cl39 Downloaded from www.physiology.org/journal/ajpcell by ${individualUser.givenNames} ${individualUser.surname} (130.070.008.131) on January 14, 2019.
Cl40
B. S. KINGSLEY
in the presence and absence of excess zinc is presented in Fig. 1. Excess zinc significantly reduced the clearance of cadmium as indicated by the reduced slope of the clearance curve. To quantify this effect, the normalized clearance (NCL) was computed. The NCL is defined as NCL=-
1 dM(t)/dt w
C(t)
t=O
where W is the liver weight (g), C(t) is the concentration of cadmium in the perfusion medium @g/ml) at time t, and dM(t)/dt is the net rate of cadmium transport (pg/ min) from the perfusion medium into the liver at time t. As defined, the NCL is the initial clearance normalized by the liver weight. In the competition experiments, the presence of excess zinc (Zn:Cd molar ratio, 13.0) decreased the NCL from the control value of 0.340 -+ 0.010 (ml/min)/g to 0.138 -+ 0.017 (ml/min)/g. The kinetics of zinc in the perfusion medium are given in Fig. 2. The isolated perfused rat liver secreted zinc into the perfusion medium at a constant rate during normal zinc control experiments; this confirmed previously reported observations (4). In the presence of excess zinc (no cadmium present) small quantities of zinc were accumulated by the liver during the 4-h experimental period. The accumulated zinc increased hepatic zinc concentrations by approximately 4 pg/g, from 34.8 t 0.1 pg/ g wet wt in normal zinc controls to 38.9 t 1.8 pg/g in excess zinc controls. When this increase in hepatic concentrations was converted into mass transferred from perfusion medium to the liver, the resulting decrease in total zinc in the perfusion medium (approximately 45 pg or 1.7% of total zinc in perfusion medium) was insignificant in the presence of measurement variability. The introduction of cadmium into the perfusion medium during normal zinc plus cadmium experiments resulted in an increase in zinc secretion rate into the perfusion medium, again confirming previous observations (4). This secretion of zinc was reflected in the decrease in hepatic zinc concentrations from 34.8 t 0.1 pg/g wet wt in normal
TIME
(HOURS)
FIG. 1. Kinetics of total cadmium in perfusion of normal zinc concentrations (A), Zn:Cd molar presence of excess zinc (B), Zn:Cd molar ratio of introduced into system at L = 0. Means -+ SE, n =
4.
J. M,
FRAZIER
zinc controls to 27.0 t 1.4 pg/g in the normal zinc plus cadmium group. The presence of cadmium in excess zinc plus cadmium experiments suppressed the slight uptake of zinc observed in excess zinc controls, and hepatic zinc concentrations increased from control levels to only 36,2 t 0.6 pg/g wet wt in this group. Biliary excretion uf cadmium and zinc. The biliary excretion of cadmium and zinc depends on both the concentrations of the metal in bile and the rate of bile formation. The effects of experimental treatments on bile formation are presented in Fig. 3, In normal zinc control experiments bile flow was relatively constant during the first half of the experiment and then dropped slightly to 83% of control rate by the last hour. Excess zinc depressed bile formation less than 10% during the first 2 h of the experimental period. By the last hour, bile formation rate dropped to 54% of control. Cadmium exposure had a significant and immediate effect on bile formation. During the 1st h of cadmium exposure in the presence of
7 2400 N
200
! -1
I1 1
0
TIME
m 1 2 (HOURS)
Im 3
I 4
FIG. 2. Kinetics of total zinc in perfusion medium in normal zinc control experiments (A), excess-zinc controls (B), normal zinc plus cadmium (C), and excess zinc plus cadmium (D). Means t SE, n = 4.
TIME medium in presence ratio of 1.6, and in 13.0. Cadmium was
AND
(HOURS)
FIG. 3. Bile flow, expressed as percent of control rate during 1st h of each experiment in normal zinc control experiment (A), excess zinc controls (B), normal zinc plus cadmium (C), and excess zinc plus cadmium (D). Means -t SE, IX = 4.
Downloaded from www.physiology.org/journal/ajpcell by ${individualUser.givenNames} ${individualUser.surname} (130.070.008.131) on January 14, 2019.
CADMIUM
TRANSPORT
IN
ISOLATED
PERFUSED
RAT
Cl41
LIVER
normal zinc, bile formation decreased to 82% of control and continued to decrease to only 18% of control at 4 h of exposure, The presence of excess zinc bad a slight protective effect. The concentration, the cumulative excretion, and the computed average hourly rate of cadmium excretion in bile are presented in Fig. 4. Cadmium rapidly appeared in bile both in the presence and absence of excess zinc. The amount of cadmium excreted in bile was reduced in the presence of excess zinc. This was predominantly due to the reduced concentration of cadmium in the bile, which in turn was related to the reduced hepatic cadmium concentration in the presence of excess zinc. Cadmium excretion rates in both treatment groups decreased
25T i
rapidly during the 3rd and 4th h due to the reduction in bile flow. The total amount of cadmium excreted during the experiment is given in Table 1 and is expressed as total micrograms and as percentage of dose taken up by the liver. Both the total cadmium excreted and the fraction of the dose taken up that was subsequently excreted into bile tended to decrease in the presence of excess zinc in the perfusion medium, but this trend was not statistically significant. The concentration, the cumulative excretion, and the computed average hourly rate of zinc excretion in bile are presented in Fig. 5. Under normal zinc control conditions, little zinc was excreted in bile during the 4-h period corresponding to cadmium exposure, Table 1. All treatments significantly increased biliary excretion of zinc. In the presence of normal zinc in perfusion medium, cadmium exposure increased biliary excretion of zinc by 3.8 lug (normal Zn plus cadmium-normal Zn control). In the presence of excess zinc, cadmium exposure increased biliary excretion of zinc by only 2.0 lug (excess Zn plus cadmium-excess Zn control). The reduction in biliary excretion of zinc induced by cadmium exposure was roughly proportional to the reduction in uptake of cadmium in the presence of excess zinc. DISCUSSION
TtME
TtME
&tOURS~
(HOURS)
The presence of excess zinc (Zn:Cd molar ratio, of 13.0) in the perfusion medium suppressed cadmium uptake by 60%. These results clearly support the hypothesis that at least one component of cadmium transport across the sinusoidal membrane of hepatocytes involves a mediated transport mechanism that is also utilized for zinc transport. A most unexpected observation in these experiments was the relatively small amount of zinc accumulated by the liver in the excess zinc control experiments. Based on the uptake of cadmium from an equivalent dose, approximately 50% of the zinc would be expected to be accumulated in the liver under these conditions. To interpret this observation, consider Table 2, where the distribution of zinc among the various compartments in the isolated perfused liver system is given under various conditions. In this table the distribution of zinc between the free species and that bound to bovine serum albumin (BSA) in the perfusion medium was computed from independent data for zinc binding to BSA at pH 7.4 and 37°C (unpublished data). In the presence of normal zinc concentrations in the perfusion medium (0.72 pg G/ml), the majority of extracellular zinc is bound to BSA, resulting TABLE I, Cumulatiue biliary excretion of cadmium and zinc in isolated perfused liver preparation Cumulative Cd Excreted in Bile, PC
TIME 4. Cadmium concentration, and average hourly rate of cadmium plus cadmium (A) and excess zinc Means t SE, n = 4. FIG.
WtOURS~ cumulative cadmium excretion, excretion in bile for normal zinc plus cadmium (B) experiments.
Normal Excess Normal Excess
Zn control Zn control Zn plus Cd Zn plus Cd
Percent of Accumulated Cd Dose Excreted
-
Cumulative %I Excreted in Bile, pg*
0.42 t 0.05 2.38 t 1.09-f
9.2 f 2.7 3.6 4: 1.3
Values are means k SE. perimental period (n = 4). control (P < 0.05).
3.3 t 0.9 1.8 t 0.6
4.23 k Lot 4.39 k 1.2-t
* Total metal excreted during 4-h ext Significantly different from normal Zn
Downloaded from www.physiology.org/journal/ajpcell by ${individualUser.givenNames} ${individualUser.surname} (130.070.008.131) on January 14, 2019.
Cl42
B. S. KINGSLEY
AND
J. M,
FRAZIER
2. Zinc distribution among various compartments of isolated perfused liver system ,, -...-.-
TABLE
I
I
I
Perfusion Bound
;XI! KE&
~
0.72
8.37 (2509)
(1084)
saturation
8.4
8.37 (2509)
8.23 (2469)
Liver Free
Free 0.002
0.002-0.023
(216)(0.6) 3.61
Complete
1
Medium
0.0232 (6.9) 0.010 (3.0)
0.0228 (6.9)
Values are concentrations in compartments, in of zinc in compartments (pg) is given in parentheses. species in perfusion medium was calculated as the zinc to BSA at pH 7.4 and 37°C. Zinc distribution water content of 80% and 58% of intracellular zinc
25.2
0.002-0.023
(0.016-O. 18) 0.010 (0.08) 0.0232
Total
34.8
25.2
34.8
(202)
(146) (348)
129 (1031)
(7463(1777)
25.2
178
34.8
cm.9 (146) (348)
(0.18)
perfused
S. KINGSLEY AND JOHN M. FRAZIER of Environmental Health Sciences, Johns Health, Baltimore, Maryland 21205
KINGSLEY, BEVERLY S., AND JOHN M. FRAZIER. Cadmium transport in isolated perfused rat Liver: zinc-cadmium competition. Am. J. Physiol, 236(3): C139-C143, 1979 or Am. J. Physiol.: Cell Physiol. 5(2): C139-C143, 1979.-The hypothesis that one component of cadmium uptake by rat hepatocytes involves a mediated transport pathway normally operative for zinc transport was tested in the isolated perfused rat liver preparation. Excess zinc in the perfusion medium suppressed cadmium uptake as indicated by the decrease in the normalized clearance (initial clearance divided by liver weight) from 0.340 t 0.019 (ml/min)/g in the presence of normal zinc concentrations (Zn:Cd molar ratio, 1.6) to 0.138 t 0.017 (ml/min)/g (Zn: Cd molar ratio, 13.0). In excess-zinc control experiments (no cadmium present) little zinc is accumulated by the liver, apparently due to competition between intrahepatic and extracellular binding. Exposure to cadmium increases both zinc secretion into the perfusion medium and biliary excretion of zinc. The effect at the sinusoidal membrane is probably a result of both the blockage of zinc resorption during cadmium uptake and the displacement of intrahepatic zinc. The effect on biliary excretion of zinc is due solely to displacement of intrahepatic zinc. These results are consistent with the proposed hypothesis for cadmium transport.
Hopkins
MATERIALS
AND
rat liver:
School
of Hygiene
METHODS
The recirculating isolated perfused rat liver preparation of Miller (6) was employed for these experiments. Male
Wistar
rats of 340 g average
weight
were
used
as
liver donors. Details of the surgical procedure were previously reported (4). The perfusion medium consisted of a Krebs-Ringer bicarbonate buffer supplemented with dextrose (0.8 g/l) and bovine serum albumin (4% wt/vol; fraction V; Sigma Chemical Corp., St. Louis, MO). Bovine serum albumin was carefully selected because the commercial product is often contaminated with zinc, copper, and iron. All experiments were performed with an initial zinc concentration of 0,66-0.90 lug Zn/ml, which approximates the normal labile serum zinc concentration in these rats determined in this laboratory. Initial volume of the perfusion medium was 350 ml and perfusion pressure was maintained at 17.5 cmHz0, corresponding to a perfusion rate of 60-75 ml/min in control experiments. The pH of the perfusion medium was maintained between 7.35-7.45 by additions of sodium bicarbonate. Each 5-h experiment was performed according to the following protocol. The first 60 min served as a control biliary excretion; mediated transport period. At 60 min (t = 0) four different treatments were imposed (four liver preparations per treatment group): 1) normal zinc control (no additions), 2) excess zinc control HEPATXC TRANSPORT OF CADMIUM is controlled by the (addition of zinc, as ZnSOd, to perfusion medium, resulting in a total zinc concentration of 8.4 lug Zn/ml or 129 following processes: 1) the transport of cadmium across FM), 3) normal zinc plus cadmium (addition of cadmium, the sinusoidal membrane of the hepatocytes, 2) intracelas CdClz, giving a concentration of 1.0 lug Cd/ml or 8.94 lular binding to cell organelles and cytoplasmic macromolecules, and 3) transport across the membrane sepa- PM and a Zn:Cd molar ratio of 1,6), and 4) excess zinc (addition of both zinc and cadmium rerating the hepatocyte from the biliary caraliculi. These plus cadmium sulting in Zn:Cd molar ratio of 13.0). At fixed intervals processes are of particular importance because the liver periods, samplays a central role in the distribution and excretion of during both the control and experimental ples of perfusion medium were collected for cadmium cadmium (5). Previous studies in this laboratory (4) have characterized cadmium-uptake kinetics in the isolated and zinc analysis. Cumulative samples of bile over l-h perfused rat liver. These studies suggested that both intervals were also collected throughout the experiment simple diffusion and mediated transport are involved in and analyzed for cadmium and zinc. Metal analyses of perfusion medium, bile, and liver cadmium transport into the hepatocytes at the sinusoidal tissue were performed by atomic absorption spectrophomembrane and that the mediated transport component may involve a pathway normally utilized for zinc trans- tometry (AA-5, Varian Techtron, Victoria, Australia) as port. The present investigation was designed to test this previously described (4). Data are reported as X t SE, rz = 4, unless otherwise indicated. Comparisons for statishypothesis in the isolated perfused rat liver preparation. If cadmium and zinc use the same membrane carrier for tical significance were performed by Student’s t test with a level of significance of P 5 0,05. uptake, then excess extracellular zinc should inhibit cadmium uptake by competing for the carrier on the sinusRESULTS oidal side of the hepatocyte membrane. Observations of the kinetics of cadmium and zinc in the perfusion medium Kinetics of cadmium and zinc in perfusion medium. and the biliary excretion of these two metals are reported. The clearance of cadmium from the perfusion medium 0363-6143/79/0000-OUOO$O1.25
Copyright 0 1979 the American Physiological Society Cl39 Downloaded from www.physiology.org/journal/ajpcell by ${individualUser.givenNames} ${individualUser.surname} (130.070.008.131) on January 14, 2019.
Cl40
B. S. KINGSLEY
in the presence and absence of excess zinc is presented in Fig. 1. Excess zinc significantly reduced the clearance of cadmium as indicated by the reduced slope of the clearance curve. To quantify this effect, the normalized clearance (NCL) was computed. The NCL is defined as NCL=-
1 dM(t)/dt w
C(t)
t=O
where W is the liver weight (g), C(t) is the concentration of cadmium in the perfusion medium @g/ml) at time t, and dM(t)/dt is the net rate of cadmium transport (pg/ min) from the perfusion medium into the liver at time t. As defined, the NCL is the initial clearance normalized by the liver weight. In the competition experiments, the presence of excess zinc (Zn:Cd molar ratio, 13.0) decreased the NCL from the control value of 0.340 -+ 0.010 (ml/min)/g to 0.138 -+ 0.017 (ml/min)/g. The kinetics of zinc in the perfusion medium are given in Fig. 2. The isolated perfused rat liver secreted zinc into the perfusion medium at a constant rate during normal zinc control experiments; this confirmed previously reported observations (4). In the presence of excess zinc (no cadmium present) small quantities of zinc were accumulated by the liver during the 4-h experimental period. The accumulated zinc increased hepatic zinc concentrations by approximately 4 pg/g, from 34.8 t 0.1 pg/ g wet wt in normal zinc controls to 38.9 t 1.8 pg/g in excess zinc controls. When this increase in hepatic concentrations was converted into mass transferred from perfusion medium to the liver, the resulting decrease in total zinc in the perfusion medium (approximately 45 pg or 1.7% of total zinc in perfusion medium) was insignificant in the presence of measurement variability. The introduction of cadmium into the perfusion medium during normal zinc plus cadmium experiments resulted in an increase in zinc secretion rate into the perfusion medium, again confirming previous observations (4). This secretion of zinc was reflected in the decrease in hepatic zinc concentrations from 34.8 t 0.1 pg/g wet wt in normal
TIME
(HOURS)
FIG. 1. Kinetics of total cadmium in perfusion of normal zinc concentrations (A), Zn:Cd molar presence of excess zinc (B), Zn:Cd molar ratio of introduced into system at L = 0. Means -+ SE, n =
4.
J. M,
FRAZIER
zinc controls to 27.0 t 1.4 pg/g in the normal zinc plus cadmium group. The presence of cadmium in excess zinc plus cadmium experiments suppressed the slight uptake of zinc observed in excess zinc controls, and hepatic zinc concentrations increased from control levels to only 36,2 t 0.6 pg/g wet wt in this group. Biliary excretion uf cadmium and zinc. The biliary excretion of cadmium and zinc depends on both the concentrations of the metal in bile and the rate of bile formation. The effects of experimental treatments on bile formation are presented in Fig. 3, In normal zinc control experiments bile flow was relatively constant during the first half of the experiment and then dropped slightly to 83% of control rate by the last hour. Excess zinc depressed bile formation less than 10% during the first 2 h of the experimental period. By the last hour, bile formation rate dropped to 54% of control. Cadmium exposure had a significant and immediate effect on bile formation. During the 1st h of cadmium exposure in the presence of
7 2400 N
200
! -1
I1 1
0
TIME
m 1 2 (HOURS)
Im 3
I 4
FIG. 2. Kinetics of total zinc in perfusion medium in normal zinc control experiments (A), excess-zinc controls (B), normal zinc plus cadmium (C), and excess zinc plus cadmium (D). Means t SE, n = 4.
TIME medium in presence ratio of 1.6, and in 13.0. Cadmium was
AND
(HOURS)
FIG. 3. Bile flow, expressed as percent of control rate during 1st h of each experiment in normal zinc control experiment (A), excess zinc controls (B), normal zinc plus cadmium (C), and excess zinc plus cadmium (D). Means -t SE, IX = 4.
Downloaded from www.physiology.org/journal/ajpcell by ${individualUser.givenNames} ${individualUser.surname} (130.070.008.131) on January 14, 2019.
CADMIUM
TRANSPORT
IN
ISOLATED
PERFUSED
RAT
Cl41
LIVER
normal zinc, bile formation decreased to 82% of control and continued to decrease to only 18% of control at 4 h of exposure, The presence of excess zinc bad a slight protective effect. The concentration, the cumulative excretion, and the computed average hourly rate of cadmium excretion in bile are presented in Fig. 4. Cadmium rapidly appeared in bile both in the presence and absence of excess zinc. The amount of cadmium excreted in bile was reduced in the presence of excess zinc. This was predominantly due to the reduced concentration of cadmium in the bile, which in turn was related to the reduced hepatic cadmium concentration in the presence of excess zinc. Cadmium excretion rates in both treatment groups decreased
25T i
rapidly during the 3rd and 4th h due to the reduction in bile flow. The total amount of cadmium excreted during the experiment is given in Table 1 and is expressed as total micrograms and as percentage of dose taken up by the liver. Both the total cadmium excreted and the fraction of the dose taken up that was subsequently excreted into bile tended to decrease in the presence of excess zinc in the perfusion medium, but this trend was not statistically significant. The concentration, the cumulative excretion, and the computed average hourly rate of zinc excretion in bile are presented in Fig. 5. Under normal zinc control conditions, little zinc was excreted in bile during the 4-h period corresponding to cadmium exposure, Table 1. All treatments significantly increased biliary excretion of zinc. In the presence of normal zinc in perfusion medium, cadmium exposure increased biliary excretion of zinc by 3.8 lug (normal Zn plus cadmium-normal Zn control). In the presence of excess zinc, cadmium exposure increased biliary excretion of zinc by only 2.0 lug (excess Zn plus cadmium-excess Zn control). The reduction in biliary excretion of zinc induced by cadmium exposure was roughly proportional to the reduction in uptake of cadmium in the presence of excess zinc. DISCUSSION
TtME
TtME
&tOURS~
(HOURS)
The presence of excess zinc (Zn:Cd molar ratio, of 13.0) in the perfusion medium suppressed cadmium uptake by 60%. These results clearly support the hypothesis that at least one component of cadmium transport across the sinusoidal membrane of hepatocytes involves a mediated transport mechanism that is also utilized for zinc transport. A most unexpected observation in these experiments was the relatively small amount of zinc accumulated by the liver in the excess zinc control experiments. Based on the uptake of cadmium from an equivalent dose, approximately 50% of the zinc would be expected to be accumulated in the liver under these conditions. To interpret this observation, consider Table 2, where the distribution of zinc among the various compartments in the isolated perfused liver system is given under various conditions. In this table the distribution of zinc between the free species and that bound to bovine serum albumin (BSA) in the perfusion medium was computed from independent data for zinc binding to BSA at pH 7.4 and 37°C (unpublished data). In the presence of normal zinc concentrations in the perfusion medium (0.72 pg G/ml), the majority of extracellular zinc is bound to BSA, resulting TABLE I, Cumulatiue biliary excretion of cadmium and zinc in isolated perfused liver preparation Cumulative Cd Excreted in Bile, PC
TIME 4. Cadmium concentration, and average hourly rate of cadmium plus cadmium (A) and excess zinc Means t SE, n = 4. FIG.
WtOURS~ cumulative cadmium excretion, excretion in bile for normal zinc plus cadmium (B) experiments.
Normal Excess Normal Excess
Zn control Zn control Zn plus Cd Zn plus Cd
Percent of Accumulated Cd Dose Excreted
-
Cumulative %I Excreted in Bile, pg*
0.42 t 0.05 2.38 t 1.09-f
9.2 f 2.7 3.6 4: 1.3
Values are means k SE. perimental period (n = 4). control (P < 0.05).
3.3 t 0.9 1.8 t 0.6
4.23 k Lot 4.39 k 1.2-t
* Total metal excreted during 4-h ext Significantly different from normal Zn
Downloaded from www.physiology.org/journal/ajpcell by ${individualUser.givenNames} ${individualUser.surname} (130.070.008.131) on January 14, 2019.
Cl42
B. S. KINGSLEY
AND
J. M,
FRAZIER
2. Zinc distribution among various compartments of isolated perfused liver system ,, -...-.-
TABLE
I
I
I
Perfusion Bound
;XI! KE&
~
0.72
8.37 (2509)
(1084)
saturation
8.4
8.37 (2509)
8.23 (2469)
Liver Free
Free 0.002
0.002-0.023
(216)(0.6) 3.61
Complete
1
Medium
0.0232 (6.9) 0.010 (3.0)
0.0228 (6.9)
Values are concentrations in compartments, in of zinc in compartments (pg) is given in parentheses. species in perfusion medium was calculated as the zinc to BSA at pH 7.4 and 37°C. Zinc distribution water content of 80% and 58% of intracellular zinc
25.2
0.002-0.023
(0.016-O. 18) 0.010 (0.08) 0.0232
Total
34.8
25.2
34.8
(202)
(146) (348)
129 (1031)
(7463(1777)
25.2
178
34.8
cm.9 (146) (348)
(0.18)
