J. Phygiol. (1978), 284, pp. 229-239 With 8 text-ftgure Printed in Great Britain

229

NUCLEOSIDE AND GLUCOSE TRANSPORT IN ERYTHROCYTES FROM NEW-BORN LAMBS

BY NUALA A. MOONEY AND J. D. YOUNG From the A.R.C. Institute of Animal Physiology, Babraham, Cambridge CB2 4AT

(Received 5 April 1978) SUMMARY

1. Glucose and inosine transport by erythrocytes from new-born lambs and adult sheep were compared. Uptake of both permeants was considerably faster in the new-born. Inosine uptake by erythrocytes from nucleoside-permeable and impermeable lambs were not significantly different at birth. The difference between the two phenotypes was first apparent 30 days after birth. 2. The post-natal changes in glucose and inosine transport activity closely paralleled the progressive decrease in the percentage of fetal erythrocytes (i.e. cells containing fetal haemoglobin) in the circulation. Cell fractionation studies confirmed that the permeability changes were directly related to changes in the relative proportions of fetal and adult haemoglobin containing erythrocytes. 3. The results demonstrate that fetal cells are highly permeable to both glucose and inosine. These cells are replaced by erythrocytes which contain adult haemoglobin and which have a much lower, but still significant, glucose permeability and either low or negligible inosine transport activity depending on the genotype of the animal. 4. Inosine transport by fetal erythrocytes from both nucleoside-permeable and impermeable animals was mediated by a nucleoside transport system which had similar properties to that responsible for nucleoside transport in adult nucleosidepermeable cells. Glucose transport in both fetal and adult cells was highly stereospecific, indicating the presence of a selective transport system. 5. It is suggested that the regulatory mechanism responsible for initiating the switch from fetal to adult haemoglobin synthesis may also be responsible for the changes in glucose and nucleoside transport activity. INTRODUCTION

Erythrocytes from fetuses and new-born animals often show different membrane transport characteristics from those of adults of the same species. This applies to glucose transport in a number of species including the sheep and pig (Widdas, 1955; Augustin, Rohden & Hacker, 1967; Lee, Auvil, Grey & Smith, 1976; Zeidler, Lee & Kim, 1976), sodium and potassium transport in the dog (Lee & Miles, 1972; Miles & Lee, 1972), cattle (Israel, MacDonald, Bernstein & Rosenmann, 1972) and sheep (Ellory & Tucker, 1969; Tucker & Ellory, 1970) and calcium activated potassium transport in the sheep (Brown, Ellory, Young & Lew, 1978). Sheep are of particular

N. A. MOONEY AND J. D. YOUNG 230 interest because as adults they show genetic polymorphism in the transport of sodium and potassium (Ellory, 1977), amino acids (Young, Ellory & Tucker, 1976) and nucleosides (Young, 1978). In the case of nucleoside transport some sheep (nucleoside-permeable phenotype) have cells which possess a high affinity transport system for both purine and pyrimidine nucleosides. Other sheep (nucleoside-impermeable phenotype) lack this system. The permeability difference between the two types of animal is under the control of two allelomorphic genes (NuI and Nui) with the gene for nucleoside impermeability (NuI) behaving as if dominant to the gene for nucleoside permeability (Nui) (Jarvis & Young, 1978). Nucleoside-permeable erythrocytes have a higher ATP concentration than nucleoside-impermeable cells suggesting that the transport system participates in the energy metabolism of the erythrocyte (Young, 1978). This paper presents data on the nucleoside permeability of erythrocytes from newborn lambs of the two phenotypes. The results demonstrate that nucleoside transport is considerably faster in the new-born and identical in the two types of animal. The changes which occur in nucleoside transport after birth are compared with the loss of glucose transport activity and the disappearance of fetal haemoglobin from the circulation. METHODS

Animals Whole blood samples were obtained from lambs and adult sheep by jugular venepuncture into heparinized evacuated tubes. All animals were maintained at Babraham under standard husbandry conditions. For the nucleoside permeability studies lambs with the genotype (Nui NuW) were produced from nucleoside-permeable x permeable matings. Nucleoside-impermeable lambs (Nu'-) were obtained from nucleoside-impermeable x impermeable crosses. Phenotype classification was confirmed when the animals were 5 months old.

Materials

[U-14C]Inosine, [3H]-D-glucose and [1-14C]-L-glucose were obtained from The Radiochemical Centre, Amersham, Bucks, U.K. Adenosine, inosine, uridine, D-glucose and L-glucose were obtained from Sigma (London) Chemical Co. Ltd, Kingston-upon-Thames, Surrey, U.K. Nitrobenzylthioinosine (6-[(4-nitrobenzyl)thio]-9-,f-D-ribofuranosyl purine) was a generous gift from Professor A. R. P. Paterson, University of Alberta Cancer Research Unit, Edmonton, Canada. Dipyridamole (Persantin Injection) was obtained from Boehringer Ingelheim Ltd, Bracknell, Berks, U.K. N-dibutyl phthalate was purchased from BDH Chemicals Ltd, Poole, Dorset, U.K. Cell preparation and fractionation Erythrocytes were washed 3 times with 20 vol. 0-92% (w/v) sodium chloride. The buffy coat was discarded at this stage except when the cells were to be subsequently fractionated. To separate erythrocytes containing fetal haemoglobin from those containing adult haemoglobin, the washed cells were resuspended in saline at a haematocrit of 40% and then centrifuged for 20 min at 1600 g in 11 0 x 0-8 cm polythene tubes (Drury & Tucker, 1963; Tucker & Ellory, 1970). After removal of the supernatant and buffy coat the cell column was sectioned into 3 equal volumes. The fractionated cells were then washed once in incubation medium (140 mM-NaCl, 5 mM-KCl, 20 mM-Tris-HCl (pH 7-2 at 37 'C), 2 mM-MgCl2 and 0.1 mM-EDTA). Unfractionated cells were also washed once in this medium. In this paper 'fetal erythrocytes' refers to erythrocytes obtained from newborn lambs and which were known to contain fetal haemoglobin. 'Adult erythrocytes' are erythrocytes obtained from either lambs or adult sheep and which contain adult haemoglobin.

PERMEABILITY OF LAMB ERYTHROCYTES

231

Transport studies Inosine and glucose uptake were measured at 37 00 by mixing 0-15 ml. prewarmed washed cells (haematocrit approximately 20% in incubation medium) with 0-15 ml. prewarmed medium containing the appropriate concentration of radioactive permeant (0-1-0.5 #sc/#mole). Incubations were stopped at predetermined time intervals (15 sec-S min) by transferring 0- 1 ml. of the cell suspension to an Eppendorf microoentrifuge tube (volume 1-5 ml.) containing 0-8 ml. ice-cold incubation medium layered on top of 0 5 ml. ice-cold n-dibutyl phthalate. The tube was immediately centrifuged at 15,000 g for 15 see using an Eppendorf 3200 microcentrifuge. The aqueous medium and dibutyl phthalate layers were removed by suction, leaving the cell pellet at the bottom of the tube. After carefully wiping the inside of the centrifuge tube with tissue paper the cell pellet was lysed with 0 5 ml. 0 5 % (v/v) Triton X-100 in water and 0-5 ml. 5 % (w/v) trichloroacetic acid was added. The precipitate was removed by centrifugation (30 see, 15,000 g) and 0 9 ml. of the supernatant transferred to 7 ml. scintillation fluid (0-1 g 1,4-bis[2(5-phenyloxazolyl)]benzene (POPOP), 5 g 2,5-diphenyloxazole (PPO) and 480 ml. Triton X-100 in 11. toluene). Samples were counted for radioactivity in a Packard Tricarb scintillation counter with quench correction. Blank values were obtained by processing cell samples which had been mixed with radioactive permeant at 0 0C. Transport rates (mole or m-mole/l. cells per hr) were calculated after subtraction of these blanks (typically < 5 % of the transport rate). In some experiments an alternative method of separating cells from extracellular medium was used. Here, incubations were stopped by the addition of 1 ml. ice-cold medium to the 0 3 ml. cell suspension, and the cells rapidly washed 4 times with 1 ml. ice-cold medium (10 sec, 15,000 g)). The washed cell pellets were treated as above. The washing procedure did not result in loss of radioactivity from the cells, and both the washing and dibutyl phthalate methods gave equivalent uptake rates (Young, 1978). Haematological measurements The hemoglobin content of cell suspensions was determined by absorbance measurements of dilute lysates at 540 nm and correlated with the packed cell volume using an experimentally determined extinction coefficient for packed erythrocytes of 290 cm-'. There was no significant difference in the value of this coefficient for fetal and adult cells or for nucleoside-permeable and impermeable erythrocytes from adult sheep (see also Upeott, Hebert & Robins, 1971). Fetal cell counts were performed using the acid elution technique of Moore, Godley, von Vliet, Lewis, Boyd & Huisman (1966). A small volume of 20 % cell suspension (0.2 ml.) was centrifuged at 15,000 g for 10 sec, and the supernatant removed. The cells were resuspended in 50 Ml. of their own plasma and a thin cell smear prepared. The slide was immediately fixed by immersion in 84% (v/v) ethanol for 5 min and then washed in water for 30 sec. Adult hemoglobin was eluted from the cells by incubation of the slide in a solution containing 0-1 M-citric acid and 0-2 M-disodium orthophosphate (pH 3 3) for 6 min at 37 'C. The slides were again washed in distilled water for 30 sec, stained in Ehrlich's acid haematoxylin for 30 min, and washed again in distilled water for a further 30 sec. The slide was finally stained for 3 min in 0 1 % (w/v) erythrosin, given a 30 sec wash in distilled water and allowed to dry. Fetal cells retained their haemoglobin under these conditions and stained strongly whereas adult cells appeared as pale ghosts. Unlike the situation in man (Wood, 1976), cells with intermediate staining were not usually seen and it was therefore possible to determine accurately the relative numbers of adult and fetal cells. Cell counts obtained by acid elution correlate closely with the percentages of adult and fetal hemoglobin estimated by starch gel electrophoresis (E. M. Tucker, personal communication). Reticulocytes were stained supravitally with brilliant cresyl blue (Archer, 1965). RESULTS

Glucose and inosine transport in erythrocytes from new-born kambs Fig. 1 shows the time-course of D-glucose (5 mM) uptake by erythrocytes from a new-born lamb compared with cells from an adult sheep. In agreement with the results of Widdas (1955) the lamb cells were considerably more permeable to glucose

232 N. A. MOONEY AND J. D. YOUNG than the adult erythrocytes. Fig. 2 compares the uptake of inosine (5 mM) by cells from a new-born lamb of the genotype Nut Nui (nucleoside-permeable) with erythrocytes from adult animals of both nucleoside-permeability types. As in the case of

2-0

2;0

/-

E

|

10

20 Time (min)

30

40

Fig. 1. D-GluCose uptake by erythrocytes from a new-born lamb and an adult sheep. The glucose concentration was 5 mm. *, Lamb erythrocytes; 0, adult sheep erythrocytes. .2 05-7 2-1

~0

~

0

20

3

5.0

7-5

0

E 1-4

E (C

00 0

2-5

10

Time (min)

Fig. 2. Inosine uptake by erythrocytes from a new-born lamb and adult nucleosidepermeable and impermeable sheep. The inosine concentration was 5 mm. *, Lamb erythrocytes; 0, adult nucleoside-permeable erythrocytes; D-1 adult nucleosideimpermeable erythrocytes.

glucose, inosine transport

was considerably faster in the lamb cells. Table 1 summarizes the initial rates Of D-glucose and inosine uptake (both 5 mm~) by erythrocytes from a number of new-born lambs and compares these rates with those obtained from cells when the same animals were 5 months old. For glucose there was a 560-fold

PERMEABILITY OF LAMB ER YTHROCT YES 233 decrease in transport activity during this period, but the cells obtained at 5 months still showed significant glucose transport activity. Inosine uptake by erythrocytes from nucleoside-permeable (Nui Nui) and impermeable lambs (NuI-) were not significantly different at birth, but had reached typical adult values by the age of 5 months. In the case of nucleoside-permeable animals there was a tenfold drop in transport activity compared with a 260-fold decrease for nucleoside-impermeable lambs. TABLE 1. Glucose and inosine uptake by erythrocytes from new-born lambs and adult sheep

Initial uptake rate

(m-mole/i. D-glUOSe (5 MM)

( Nuoleoside-permeable Inosine (5 mM)

(Nu NWu)

Nueleoside-impermeable (Nur-)

Age 1 day 5 months 2 days 5 months 2 days 5 months

No. of

Ratio animals 3 560

cells per hr) 510+18 091 + 0-08 20X2+0*8 1*93 ± 0.16

10.5

185+±1.4

264

6 4

0-07 ± 0'04

Experimental details are described in the text. Values are means + s.E. of means.

75 In

03 o 0-. 0-4

oL'

50 4-

-

aR 0.

0

Y

0*1 10

20

30

40

10

50

20

30

40

Days after birth

Fig. 3. Post-natal changes in erythrocyte D-glucose permeability and fetal cell count. The glucose concentration was 5 mM. Values are the means ( i.E. of means) for three animals.

Change in glucose and nucleoide transport with age Figs. 3 and 4 show in detail the age-related changes in glucose and inosine transport (5 mM) and compare the loss of transport activity with the time course of disappearance of fetal cells from the circulation. In all cases there was a rapid and parallel decrease of transport activity and fetal cell count during the first 50 days after birth. In the case of inosine transport, animals of the two phenotypes could be readily distinguished by 40 days after birth. Fig. 5 shows the correlation between glucose and inosine transport rates and the

N. A. MOONEY AND J. D. YOUNG 234 fetal cell count. For both permeants there was a highly significant linear correlation between the two parameters (correlation coefficients 0 995, 0-988 and 0-998 for glucose transport, inosine transport by nucleoside-permeable animals and inosine uptake by nucleoside-impermeable animals respectively). Linear regression analyses 24

100

18

75

01

-iU

0 U

Cu 50

-' 12

4-

0

E

25 L 0.

10

20

30

40

50

60

10

20

30

40

50

60

Days after birth

Fig. 4. Post-natal changes in erythrocyte inosine permeability and fetal cell count in nucleoside-permeable and impermeable lambs. The inosine concentration was 5 mM. Values are the means (± sx.. of means) for four nucleoside-impermeable (0) and six nuoleoside-permeable (0) animals. 100 75 2

50 40>

25

5

10

15

20

25

200

400

600

Uptake (m-mole/l. cells per hr)

Fig. 5. Correlation between post-natal erythrocyte glucose and inosine permeability changes and fetal cell count. The transport rates and fetal cell counts ( ± s.x. of means) are taken from Figs. 3 and 4. A, inosine transport by nucleoside-permeable (@) and impermeable (0) lambs. B, D-glucose transport. The parameters of the linear regression lines are given in the text.

of the data gave intercepts (+ s.E.) of 43 + 15, 4-33 + 0-79 and 0-22 + 0-35 m-mole/l. cells per hr for glucose uptake, inosine uptake by nucleoside-permeable animals and inosine transport by nucleoside-impermeable lambs respectively, with gradients (+ S.E.) of 4-89 + 0-28, 0-163 + 0-015 and 0-189 + 0-007 respectively. These results strongly suggest that the loss of glucose and inosine transport activity is a direct consequence of the loss and/or dilution of fetal erythrocytes and

235 PERMEABILITY OF LAMB ERYTHROCYTES the appearance of adult cells in the circulation. The relationship between transport activity and fetal cell count was further investigated by fractionating cell samples into three equal volumes according to cell density. Fig. 6 shows the results obtained for glucose permeability, and demonstrates that in agreement with previous results 100

06 fAd;S

75 a)

04

-

m

4-

50

a)

- 023

25

D 0-1 10

20

50 Days after birth

10

20

30

40

50

Fig. 6. D-Glucose permeabilities and fetal cell counts of fractionated lamb erythrocytes. Erythrocytes were fractionated into three equal volumes as described in the text. @, Fraction 1 (top fraction); 0O fraction 2 (middle fraction); *, fraction 3 (bottom fraction). The glucose concentration was 5 mm. Values are the means ± (9.E. of means) for three animals. 24 -c a)

[

W

-.

a) C.

12

-a a1)

No-

E E

oe 6

0.1 :D I

I

I

I

I

I

10

20

30

40

50

60

Days after birth

Fig. 7. Inosine permeabilities and fetal cell counts of fractionated erythrocytes from nucleoside-permeable lambs. Erythrocytes were fractionated into three equal volumes as described in the text. *, Fraction 1 (top fraction); 0, fraction 2 (middle fraction); fraction 3 (bottom fraction). The inosine concentration was 5 mm. Values are the means ( ± s.R. of means) for six animals. *,

(Ellory & Tucker, 1969) the fractionation technique effectively separated fetal from adult erythrocytes. Fetal cells were progressively and sequentially lost from each of the three fractions, and the changes in glucose transport activity of the three

236 N.A. MOONEY AND J. D. YOUNG fractions closely paralleled the loss of fetal cells from the same fractions. Similar results were obtained for inosine transport in both nucleoside-permeable (Fig. 7) and nucleoside-impermeable (Fig. 8) lambs. Reticulocytes were often present in the top fraction (fraction 1) but never exceeded 3% of the cell population. No reticulocytes were found in the other two fractions. Kinetic and inhibitor studies For this series of experiments fetal erythrocytes were harvested from blood samples obtained from 3-week-old nucleoside-permeable and impermeable lambs. The fetal cell preparations used in these experiments contained > 75% fetal cells and were obtained by the separation technique described in the Methods section.

CL

0

20

100

15

75

10

20

-

50

E

4a, 5

~~~

~

~~~~~~~25-T

Q.

10

20

30

40

50

60

10

20

30

40

50

60

Days after birth

Fig. 8. Inosine permeabilities and fetal oell counts of fractionated erythrocytes from nucleoside-impermeable lambs. Erythrocytes were fractionated into three equal volumes as described in the text. *, Fraction 1 (top fraction); 0, fraction 2 (middle fraction); N, fraction 3 (bottom fraction). The inosine concentration was 5 mm. Values are the means ( ± s.E. of means) for four animals.

The concentration dependence of inosine uptake by fetal cells from nucleosidepermeable and impermeable lambs was saturable. In both cases uptake was consistent with simple Michaelis-Menten kinetics giving apparent Km values of 0 35 and 0-4 mM for the nucleoside-permeable and impermeable lambs respectively. For comparison, the apparent Km value for inosine uptake by nucleoside-permeable cells from adult sheep has previously been estimated to be 0-26 mm (Young, 1978). Inosine uptake by nucleoside-impermeable erythrocytes from adult sheep is slow and nonsaturable. A number of vasodilator drugs and S-substituted-6-thiopurine ribonucleosides are potent inhibitors of nucleoside transport in a variety of cell types (see for example Berlin & Oliver, 1975). Both dipyridamole and nitrobenzylthioinosine were extremely effective inhibitors of inosine uptake (1 mM) by fetal cells from both types of animal (> 80% inhibition at 1 /SM). Uridine and adenosine (5 mM) were also effective inhibitors of inosine uptake (1 mM) by fetal cells from nucleoside-permeable and impermeable animals (65% and 78% inhibition respectively with uridine and

237 PERMEABILITY OF LAMB ERYTHROCYTES 73 and 79% inhibition respectively with adenosine). The degree of inhibition in all cases was similar to that given by adult permeable erythrocytes (see Young, 1978). The kinetics of glucose transport in fetal and adult erythrocytes were not investigated in detail. However, transport in both cell types was highly stereospecific

(> 95% at5mM). DISCUSSION

The present results confirm and extend the initial study of Widdas (1955) which demonstrated that erythrocytes from new-born lambs transport glucose considerably faster than erythrocytes from adult sheep. The present study also shows that inosine transport is much more rapid in the new-born. Furthermore erythrocytes from potentially nucleoside-permeable and impermeable animals show indistinguishable transport rates at this stage. The difference between the two types of animal is first apparent approximately 30 days after birth. Similar post-natal changes in glucose permeability have been observed in a number of other species including the pig (Widdas, 1955; Augustin et al. 1967; Lee et al. 1976). Kim & Luthra (1977) using density separation and 5l0r and 59Fe labelling techniques suggested that the loss of transport activity in the pig after birth resulted from the replacement of glucose-permeable cells with glucose-impermeable erythrocytes. Unlike the pig, the sheep possesses a distinct fetal haemoglobin, and in the present series of experiments it was possible to compare directly the changes in the two parameters. The observed decrease in both glucose and inosine transport activity after birth closely paralleled the progressive decrease in the percentage of fetal erythrocytes (i.e. cells containing fetal haemoglobin) in the circulation. Indeed the experimental results (Fig. 5) suggest that the observed uptake rate (v) during the post-natal period was simply related to the relative proportions of fetal (f) and adult (1 -f) cells in the following way v = f.a+ (1 -f).b, where a represents the transport rate of the fetal cell and b that of the adult cell. In the case of glucose transport the permeability of the adult cells appearing after birth is twelvefold lower than the permeability of the fetal cells. Similarly, in the case of inosine uptake in nucleoside-permeable animals there is a fivefold permeability difference between the two cell types. In the case of nucleoside-impermeable animals the fetal cells are replaced by erythrocytes which are impermeable to inosine. The glucose permeability of the adult cells entering the circulation after birth is significantly higher than that of adult cells from older animals (Fig. 5 and Table 1). A similar, though less marked difference is also apparent for inosine transport in the case of nucleoside-permeable animals. The most likely explanation is that this is a reflexion of the difference in mean cell age of adult erythrocytes from young lambs and adult sheep. Preliminary experiments suggest that reticulocytes from adult sheep have higher glucose and inosine permeabilities than mature erythrocytes (J. D. Young, unpublished observations). Inosine transport by fetal erythrocytes from both nucleoside-permeable and impermeable animals was saturable and inhibited by micromolar concentrations of dipyridamole and nitrobenzylthioinosine. Uridine and adenosine were also effective inhibitors of inosine uptake. Thus inosine transport in these cells is mediated by a

N. A. MOONEY AND J. D. YOUNG nucleoside-transport system which has similar properties to that responsible for nucleoside transport in adult nucleoside-permeable erythrocytes. Although the kinetics of glucose transport were not investigated in detail, it was demonstrated that transport in both fetal and adult cells was highly stereospecific suggesting the presence of a selective transport system rather than uptake by simple diffusion (see also Widdas, 1955; Zeidler et al. 1976). The control mechanism responsible for the transition from fetal to adult haemoglobin is not well understood either at the cellular or molecular level, and in particular the factor(s) responsible for initiating the switch have not been identified (see for example Wood, 1976). Nevertheless it is clear that the transition is a consequence of selective gene regulation (Nigon & Godet, 1976). Since the transport changes described in this paper occur coincidentally with the switch from fetal to adult haemoglobin, it is possible that the same regulatory mechanism may initiate both the membrane permeability and haemoglobin changes. Other erythrocyte transport changes which occur at the same time in this species include the calcium-activated potassium channel (Brown et al. 1978) and potassium transport through the sodium pump (Ellory & Tucker, 1969). The calcium-activated potassium channel is present in the new-born lamb but absent from adult sheep. Potassium transport through the sodium pump is of particular interest because adult sheep show a genetic polymorphism, giving rise to high potassium and low potassium phenotypes (see for example Ellory, 1977). At birth low potassium lambs have high erythrocyte potassium concentrations and, as is the case for nucleoside transport, the difference between high and low potassium animals is only apparent some 30 days after birth. In marked contrast, amino acid transport in the erythrocytes of new-born lambs is very similar to that in adult sheep and a genetic lesion in adult sheep resulting in the functional absence of the major amino acid transport system is also expressed at birth (Young et al. 1978). The changes in transport activity after birth, although widespread, are therefore selective. The presence of fetal haemoglobin with its high oxygen affinity is physiologically advantageous in facilitating oxygen transport across the placenta. The distinctive membrane permeability characteristics of fetal sheep erythrocytes may be of equal significance during fetal development, and further investigation of the changes occurring in transport systems and their genetic variants during post-natal development may help elucidate both the regulatory mechanisms responsible for the permeability changes and their physiological significance. 238

REFERENCES

ARCHER, R. K. (1965). Haematological Techniques for U8e on Animals, pp. 75-76. Oxford: Blackwell Scientific Publications. AUGUSTIN, H. W., ROHDEN, L. V. & HACKER, M. R. (1967). tVber einige Eigenschaften des monosaccharid Transportsystems in Erythrozyten neugeborenen und erwachsener Kaninchen. Acta biol. med. germ. 19, 723-735. BERLN, R. D. & OLIVER, J. M. (1975). Membrane transport of purine and pyrimidine bases and nucleosides in animal cells. Int. Rev. Cytol. 42, 49-101. BROWN, A. M., EMLORY, J. C., YOUNG, J. D. & LEW, V. L. (1978). A calcium activated potassium channel present in foetal red cells of the sheep but absent from reticulocytes and adult red cells. Biochim. biophy8. Acta 511, 163-175.

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DRuRy, A. N. & TucKER, E. M. (1963). Red cell volume, potassium and haemoglobin changes in lambs. Res. vet Sci. 4, 568-579. ELLORY, J. C. (1977). The sodium pump in ruminant red cells. In Membrane Transport in Red Cells, ed. ELLORY, J. C. & LEW, V. L., pp. 363-382. London: Academic Press. ELLORY, J. C. & TUCKER, E. M. (1969). Active potassium transport and the development of m antigen on the red cells of LK type lambs. J. PhyIiol. 204, 101P. ISRAEL, Y., MACDONALD, A., BERNSTEIN, J. & RosENMANN, E. (1972). Changes from high potassium (HK) to low potassium (LK) in bovine red cells. J. gen. Physiol. 59, 270-284. JARVIS, S. M. & YOUNG, J. D. (1978). The genetic control of nucleoside transport variation in sheep erythrocytes. Biochem. Genet. (In the Press). KIM, H. D. & LUTHRA, M. G. (1977). Pig reticulocytes. III. Glucose permeability in naturally occurring reticulocytes and red cells from newborn piglets. J. gen. Physiol. 70, 171-185. LEE, P. & MiiiTs, P. R. (1972). Density distribution and cation composition of red blood cells in newborn puppies. J. cell. comp. Phfysiol. 79, 377-388. LEE, P., AUVIL, J., GREY, J. E. & SMITH, M. (1976). 3-0-Methyl glucose transport in newborn and adult dog red cells. Fedn Proc. 35, 780. MiLEs, P. & LEE, P. (1972). Sodium and potassium content and membrane transport properties in red blood cells from newborn puppies. J. cell. comp. Physiol. 79, 367-376. MOORE, S. L., GODLEY, W. C., VON VLIT, G., LEwIs, J. P., BOYD, E. & Huism", T. H. J. (1966). The production of haemoglobin C in sheep carrying the gene for haemoglobin A: haematologic aspects. Blood 28, 314-329. NIGON, V. & GODET, J. (1976). Genetic and morphogenetic factors in haemoglobin synthesis during higher vertebrate development: an approach to cell differentiation mechanisms. Int. Rev. Cytol. 46, 79-176. TUCKER, E. M. & ELLORY, J. C. (1970). The M-L blood group system and its influence on red cell potassium levels in sheep. Anim. Blood Groups & Biochem. Genet. 1, 101-112. UPcowT, D. H., HEBERT, C. N. & RoBINs, M. (1971). Erythrocytes and leukocyte parameters in newborn lambs. Re8. vet. Sci. 12, 474-477. WIDDAS, W. F. (1955). Hexose permeability of foetal erythrocytes. J. Physiol. 127, 318-327. WOOD W. G. (1976). Haemoglobin synthesis during human foetal development. Br. med. Bull. 32, 282-287. YOUNG, J. D. (1978). Nucleoside transport in sheep erythrocytes: genetically controlled transport variation and its influence on erythrocyte ATP concentrations. J. Physiol. 277, 325-339. YOUNG, J. D., ELLORY, J. C. & TUCKER, E. M. (1976). Amino acid transport in normal and glutathione-deficient sheep erythrocytes. Biochem. J. 154, 43-48. YOUNG, J. D., ELLORY, J. C. & TUCKE1R, E. M. (1978). Amino acid transport properties of erythrocytes from normal newborn lambs and lambs with an inherited defect in amino acid transport. Biochem. biophys. Acta. (In the Press). ZEIDLER R. B. LEE, P. & KIM, H. D. (1976). Kinetics of 3-0-methyl glucose transport in red blood cells of newborn pigs. J. gen. Physiol. 67, 67-80.

Nucleoside and glucose transport in erythrocytes from new-born lambs.

J. Phygiol. (1978), 284, pp. 229-239 With 8 text-ftgure Printed in Great Britain 229 NUCLEOSIDE AND GLUCOSE TRANSPORT IN ERYTHROCYTES FROM NEW-BORN...
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