SHORT COMMUNICATION Effect of Lactation on Lipolysis in Rat Adipose Tissue ABSTRACT
MATERIALS AND METHODS
The c o n c e n t r a t i o n s of nonesterified fatty acids and glycerol in rat parametrial adipose tissue increased at peak lactation. Adipose tissue f r o m lactating rats showed higher rates of release of nonesterified fatty acids and glycerol when incubated in vitro than did tissue f r o m nonlactating rats, but there was a substantial increase in the esterification o f fatty acids during involution. These results support earlier evidence that fat reserves were mobilized during lactation.
H o o d e d Norway rats f r o m the Institute c o l o n y were used and were 3 m o n t h s old at time of mating. Details of their m a n a g e m e n t and feeding were as described previously (4). In addition to a group of u n m a t e d animals, groups of rats were studied on the 2nd and 14th days of lactation and the 3rd day after removing the young. In the last group, the y o u n g were r e m o v e d on the 21st day after birth. The rats were killed by breaking their necks and the parametrial fat bodies were quickly exposed. To obtain an estimate of the c o n t e n t s o f n o n e s t e r i f i e d f a t t y acids ( N E F A ) and glycerol in vivo, portions of this tissue were r e m o v e d , i m m e d i a t e l y frozen in liquid nitrogen, a n d a n a l y z e d for N E F A and glycerol as described below. F u r t h e r p o r t i o n s o f tissue (200-300 mg) were rapidly weighed and placed in small E r l e n m e y e r flasks with 3 ml of a modified Krebs Ringer bicarbonate m e d i u m (pH 7.4), which c o n t a i n e d albumin (30 mg/ml) and one-half the r e c o m m e n d e d a m o u n t of calcium. Epinephrine, when used, was added to a final c o n c e n t r a t i o n of 1/~g/ml. Incubations were carried out at 37 C with c o n t i n u o u s shaking for 60 min. A t the end of the i n c u b a t i o n period, the flask c o n t e n t s were h o m o g e n i z e d and a p o r t i o n of the h o m o g e n a t e (1 ml) was used for the e x t r a c t i o n of N E F A as described by Dole and Meinertz (5). The final heptane layer which contained the N E F A was r e m o v e d , taken to dryness under nitrogen, and redissolved in chloroform. Nonesterified f a t t y acids were d e t e r m i n e d by a colorimetric m e t h o d (6). A further p o r t i o n of the h o m o g e n a t e (1.8 ml) was added to 0.2 ml o f 1 M m e t a p h o s p h o r i c acid and mixed. A f t e r centrifugation, the clear solution b e t w e e n the precipitated protein and supernatant fat was r e m o v e d , neutralized with 1 M NaOH, and used for the d e t e r m i n a t i o n of glycerol by an e n z y m a t i c m e t h o d (7). Bovine serum albumin ( F r a c t i o n V) was o b t a i n e d from BDH Chemicals Ltd. (Poole, England), and epinephrine was f r o m Evans Medical Ltd. (Liverpool, England).
INTRODUCTION
E a r l y studies based on carcass analysis d e m o n s t r a t e d that female rats retained and stored a large a m o u n t of fat during pregnancy. These additional reserves disappeared during lactation, and it was assumed that they were utilized to meet the demands of milk production (1). Subsequent w o r k showed that fat storage was confined to the first two-thirds of pregnancy (2) and revealed that the end of pregnancy was characterized by an increasing mobilization of these reserves (2,3). Recent studies in vitro in this l a b o r a t o r y (4) showed that f a t t y acid synthesis f r o m glucose by rat adipose tissue virtually ceased during lactation but increased to very high levels after the y o u n g were removed. These results provided support for the concept that lipid stores were mobilized during lactation and replaced during involution. In this c o m m u n i c a t i o n we report additional evidence in support of this conclusion. TABLE I Effect o f Lactation on Contents o f N o n e s t e r i f i e d
Fatty Acids (NEFA) and Glycerol in Rat Adipose Tissue a Group Unmated 2 days lactation 14 days lactation 3 days involution
NEFA 2.23 3.38 4.19 1.69
+- 0.29 -+ 0.54 b + 0.34 b + 0.25
Glycerol
1.03 1.33 1.70 1.43
+- 0.08 +- 0.16 -+ 0.14 b -+ 0.10 b
aResults are expressed as #mol/g of tissue and represent means _+SE for six rats.
RESULTS AND DISCUSSION
bValues significantly (P
MATERIALS AND METHODS
The c o n c e n t r a t i o n s of nonesterified fatty acids and glycerol in rat parametrial adipose tissue increased at peak lactation. Adipose tissue f r o m lactating rats showed higher rates of release of nonesterified fatty acids and glycerol when incubated in vitro than did tissue f r o m nonlactating rats, but there was a substantial increase in the esterification o f fatty acids during involution. These results support earlier evidence that fat reserves were mobilized during lactation.
H o o d e d Norway rats f r o m the Institute c o l o n y were used and were 3 m o n t h s old at time of mating. Details of their m a n a g e m e n t and feeding were as described previously (4). In addition to a group of u n m a t e d animals, groups of rats were studied on the 2nd and 14th days of lactation and the 3rd day after removing the young. In the last group, the y o u n g were r e m o v e d on the 21st day after birth. The rats were killed by breaking their necks and the parametrial fat bodies were quickly exposed. To obtain an estimate of the c o n t e n t s o f n o n e s t e r i f i e d f a t t y acids ( N E F A ) and glycerol in vivo, portions of this tissue were r e m o v e d , i m m e d i a t e l y frozen in liquid nitrogen, a n d a n a l y z e d for N E F A and glycerol as described below. F u r t h e r p o r t i o n s o f tissue (200-300 mg) were rapidly weighed and placed in small E r l e n m e y e r flasks with 3 ml of a modified Krebs Ringer bicarbonate m e d i u m (pH 7.4), which c o n t a i n e d albumin (30 mg/ml) and one-half the r e c o m m e n d e d a m o u n t of calcium. Epinephrine, when used, was added to a final c o n c e n t r a t i o n of 1/~g/ml. Incubations were carried out at 37 C with c o n t i n u o u s shaking for 60 min. A t the end of the i n c u b a t i o n period, the flask c o n t e n t s were h o m o g e n i z e d and a p o r t i o n of the h o m o g e n a t e (1 ml) was used for the e x t r a c t i o n of N E F A as described by Dole and Meinertz (5). The final heptane layer which contained the N E F A was r e m o v e d , taken to dryness under nitrogen, and redissolved in chloroform. Nonesterified f a t t y acids were d e t e r m i n e d by a colorimetric m e t h o d (6). A further p o r t i o n of the h o m o g e n a t e (1.8 ml) was added to 0.2 ml o f 1 M m e t a p h o s p h o r i c acid and mixed. A f t e r centrifugation, the clear solution b e t w e e n the precipitated protein and supernatant fat was r e m o v e d , neutralized with 1 M NaOH, and used for the d e t e r m i n a t i o n of glycerol by an e n z y m a t i c m e t h o d (7). Bovine serum albumin ( F r a c t i o n V) was o b t a i n e d from BDH Chemicals Ltd. (Poole, England), and epinephrine was f r o m Evans Medical Ltd. (Liverpool, England).
INTRODUCTION
E a r l y studies based on carcass analysis d e m o n s t r a t e d that female rats retained and stored a large a m o u n t of fat during pregnancy. These additional reserves disappeared during lactation, and it was assumed that they were utilized to meet the demands of milk production (1). Subsequent w o r k showed that fat storage was confined to the first two-thirds of pregnancy (2) and revealed that the end of pregnancy was characterized by an increasing mobilization of these reserves (2,3). Recent studies in vitro in this l a b o r a t o r y (4) showed that f a t t y acid synthesis f r o m glucose by rat adipose tissue virtually ceased during lactation but increased to very high levels after the y o u n g were removed. These results provided support for the concept that lipid stores were mobilized during lactation and replaced during involution. In this c o m m u n i c a t i o n we report additional evidence in support of this conclusion. TABLE I Effect o f Lactation on Contents o f N o n e s t e r i f i e d
Fatty Acids (NEFA) and Glycerol in Rat Adipose Tissue a Group Unmated 2 days lactation 14 days lactation 3 days involution
NEFA 2.23 3.38 4.19 1.69
+- 0.29 -+ 0.54 b + 0.34 b + 0.25
Glycerol
1.03 1.33 1.70 1.43
+- 0.08 +- 0.16 -+ 0.14 b -+ 0.10 b
aResults are expressed as #mol/g of tissue and represent means _+SE for six rats.
RESULTS AND DISCUSSION
bValues significantly (P
