Net hepatic lactate balance following mixed meal feeding in the four-day fasted conscious dog

Net Hepatic

Lactate Balance Following Mixed Meal Feeding in the Four-Day Fasted Conscious Dog Mary A. Davis, Phillip E. Williams,

and Alan D. Cherrington

The present experiments were undertaken to determine whether four days of fasting and marked hapatic glycogen depletion would alter the effect of mixed meal feeding on net hepatic lactate balance in the conscious dog. Dogs were fasted for four days and were then fed a mixed meal over a ten-minute period. Net hepatic glucose and lactate balance were monitored for the next eight hours using the A-V difference technique. The arterial plasma glucose level rose to a maximum of 121 f 3 mg/dL three hours after feeding and then decreased. Net hepatic glucose output declined to 0.44 f 0.44 mg/kg/min but the liver never became a net consumer of glucose. The arterial blood lactate level rose from 676 * 71 to 1000 2 158 I.tmol/L as the liver switched from net lactate uptake (12.2 ? 2.0 ~mol/kg/min) to net lactate production (4.3 ? 1.7 I.rmol/kg/min). Over the course of the eight-hour postprandial period 25 g of glycogen were deposited in the liver. The net hepatic uptake of the gluconeogenic amino acids rose from 6.1 t 1.2 fimol/kg/min to a peak of 15.4 f 4.3 gmollkgglmin one hour after feeding. Net hepatic uptake of glycerol fell from 3.0 t 0.3 pmollkglmin to an average of 1.5 ? 0.4 pmol/kg/min. The plasma insulin level increased from 13 f 2 @/mL to a peak of 50 + 4 wU/mL at 3.5 hours and fell to 32 & 7 @/mL by 8 hours. The plasma glucagon level rose from 22 + 3 pg/mL to 93 + 12 pg/mL 1.5 hours after feeding and fell to 68 & 6 pg/mL 8 hours after feeding. It is concluded that in the long-fasted dog fed a mixed meal, (I 1the liver switches from net lactate uptake to net lactate output, (2) net hepatic glucose production decreases, but net hepatic glucose uptake does not occur, and (3) hepatic glycogen synthesis occurs despite the absence of net glucose and lactate uptake by the liver. EI1987 by Grune & Stratton, Inc.

T

HE LIVER of the overnight (18 hour) fasted conscious dog is known to produce rather than consume lactate.’ In addition, it has been shown that net hepatic lactate production increases more than five fold in such animals within an hour of mixed meal consumption, and that net lactate production continues for almost 24 hours following feeding.’ Nevertheless, during the first 12 postprandial hours significant quantities (44 g) of glycogen are stored in the liver.’ The dog has a marked capacity to store liver glycogen following feeding, and it remains unclear to what extent the hepatic glycogen level determines the net hepatic lactate response to feeding. In order to examine the relationship between hepatic glycogen content and the changes in hepatic lactate balance which occur following consumption of a mixed meal, we repeated our earlier study carried out on dogs faster overnight in animals which had been fasted for 96 hours. The aim of the present experiments, therefore, was to determine the effect of fasting, and thus a decrease in the hepatic glycogen content, on net hepatic lactate balance after feeding. MATERIALS

AND METHODS

Animals and Surgical Procedures Experiments were carried out on five 96-hour fasted, conscious dogs (17 to 23 kg), of either sex which had been fed a diet of meat and chow (Kal Kan meat and Wayne Lab blox; 31 percent protein,

From the Department of Molecular Physiology and Biophysics and the Department of Surgery, Vanderbilt Universiiy Medical School, Nashville. Address reprint requests to Alan D. Cherrington, PhD. Department of Molecular Physiology and Biophysics, Vanderbilt University Medical School, 2Ist Ave South at Garland Ave. Nashville, TN 37232. Q 1987 by Grune & Stratton, Inc. 0026-0495/87/3609-0008$03.00/O

856

52 percent carbohydrate, 11 percent fat, and 6 percent fiber based on dry weight, ie, an average of 129 g protein, 2 12 g carbohydrate, 48 g fat, and 24 g fiber) once daily prior to the 96-hour fast. It should be noted that the carbohydrate content of the diet was determined by the difference between the sum of the weights of protein, fat, and crude fiber in the meal and the total weight of the meal. It thus includes a significant but undefined amount of indigestible carbohydrate. In fact, after extensive acid hydrolysis the average meal was found to contain only 105 g of glucose, as much as 25% of which may have been in an indigestible form, such as cellulose, prior to acid treatment. The protein content of the meal was confirmed by amino acid analysis of an acid hydrolyzed sample of the diet, but, as with the carbohydrate, not all of the protein was necessarily in a digestible form. Sixteen days prior to each experiment a laparotomy was performed under general anesthesia (sodium pentobarbital 25 mg/kg), and silastic catheters were inserted into the portal vein and the left common hepatic vein. The tip of the portal vein catheter was placed 2 cm from the point at which the vessel enters the liver, and the tip of the hepatic vein catheter was placed 1 cm inside the left common hepatic vein. After a cutdown over the left inguinal area, an additional sampling catheter was inserted into a branch of the femoral artery. After the catheters were inserted, they were filled with saline containing heparin (200 U/mL, Abbot Laboratories, North Chicago), and their free ends were knotted and placed in subcutaneous pockets so that complete closure of both skin incisions was possible. Two weeks after surgery, blood was withdrawn to determine the hematocrit and the leukocyte count of the animal. Only animals which had: (1) a leukocyte count below 16,00O/mm’, (2) a hematocrit above 38%, (3) a good appetite (consuming all of the daily ration prior to initiation of the fast), and (4) normal stools were used. On the day of the experiment, the subcutaneous ends of the catheters were freed through a small skin incision made under local anesthesia (2% lidocaine, Astra Pharmaceutical Products, Inc. Worcester, MA). The contents of each catheter were aspirated, and catheters were flushed with saline. An Angiocath (18 gauge, Abbot) was inserted into the right cephalic vein for infusion of indocyanine green. After pre-experimental preparation, the dog was allowed to stand quietly in the Pavlov harness for 20 to 30 minutes prior to the beginning of the experiment. Metabolism, Vol 36, No 9 (September), 1987: pp 856-862

857

NET HEPATIC LACTATE BALANCE

Experimental Design Each experiment consisted of a 30-minute control period (-0.5 hours to 0 hours) and an eight hour sampling period which followed consumption (over 10 min) of the meal. The test meal (meat 18.9 g/kg, and chow 23.3 g/kg) was of the same composition as the dogs’ usual meal (see above). Blood samples were drawn from each catheter every 15 minutes during the control period, every 30 minutes during the first four hours following the meal and every hour thereafter. Indocyanine green was infused (0.10 mg/m2/min) throughout the experiment for the estimation of hepatic blood Row.

Processing of Blood Samples The collection and immediate processing of blood samples have been described previously.’ It should be noted, however, that the arterial and portal blood samples were collected simultaneously, approximately 30 seconds prior to collection of the hepatic venous samples. This was done in an attempt to compensate for the time required for the blood to pass through the liver’ and thus allow accurate estimates of hepatic balance to be made even in non-steady state conditions. Plasma glucose concentrations were determined using the glucose-oxidase method in a Beckman glucose analyzer (Beckman, Fullerton, CA). Blood lactate and glycerol concentra-

tions were determined using a Technicon autoanalyzer according to the method developed by Lloyd et al4 in samples deproteinized with 4% (w/v) perchloric acid (1 mL blood + 3 mL PCA). Amino acid concentrations in blood samples deproteinized with 10 percent (w/v) sulfosalicylic acid (1 mL blood + 1 mL SSA) were determined using a Beckman Amino Acid Analyzer as described by Bloomgarden et al.’ The concentrations of glutamine and glutamate in whole blood samples deproteinized with 4% (w/v) perchloric acid (1 mL blood + 3 mL PCA) were determined using a modification of the methods of Lund6 and of Bernt and Bergmeyer’ adapted for use on the Technicon autoanalyzer. Blood urea nitrogen levels were determined spectrophotometrically.* The hepatic glycogen content was determined in quick frozen liver slices according to the method of Chan and Exton.’ Samples were taken from each lobe of the liver of each animal in order to minimize any effects of possible differences in glycogen distribution within the liver. The glucagon concentration in plasma samples to which 500 U/mL Trasylol had been added was determined using the 30 K antiserum of Unger according to the method of Aguilar-Parada et al.” Plasma insulin was measured using the Sephadex bound antibody procedure.”

Materials

rather than plasma values of these parameters would have altered hepatic balance so that net output or uptake would have been 17% greater than the values reported in this paper (for discussion of this issue see reference 25). The proportion of the hepatic blood supply provided by the hepatic artery was assumed to be 28% based on a compilation of data from many sources by Greenway and Stark.” The changes (-20%) in hepatic blood flow which occur only during the first hour after the meal have been attributed to a selective increase in portal Row.‘~ Since in the present study, however, we did not observe any significant changes in total flow, the distribution of flow was assumed not to have changed during the postprandial period. Statistical comparisons were made using the paired t test according to Snedecor and Cochran.”

RESULTS

Pancreatic Hormones The arterial plasma insulin level rose from 13 f 2 pU/mL to a maximum of 50 + 4 pU/mL at 3.5 hours (P < .OS) and was still elevated (32 ‘- 7 rU/mL, P < .OS) 8 hours after feeding (Fig 1). Arterial plasma glucagon increased from 22 + 3 pg/mL to a maximum of 93 * 12 pg/mL (P < .05) by I.5 hours and then fell to 68 f 6 pg/mL, (P c .05) by 8 hours. Glucose Metabolism Arterial plasma glucose rose from a control period level of 91 f 3 mg/dL to a maximum level of 121 + 3 mg/dL (P < .05) at 3 hours, and was still elevated 8 hours after feeding (Fig 2). The gut switched from net glucose consumption (1.15 + 0.26 mg/kg/min) to net glucose production by 30 minutes. Net hepatic glucose output decreased slowly following feeding and was significantly reduced by the end of the experiment (0.44 + 0.44 mg/kg/min). MEAL J 60 ARTERIAL PLASMA INSULIN 4o (IrU/ml)

-

Phadebas Insulin Radioimmunoassay kits were purchased from Pharmacia Fine Chemicals, Piscataway, NJ. Trasylol was obtained from FBC Pharmaceuticals, Inc., NY. Glucagon 30K antiserum was purchased from University of Texas, Southwestern Medical School and ‘251-labeled glucagon and standard glucagon were obtained from Novo, Copenhagen. Indocyanine green was purchased from Hynson. Westcott and Dunning; Baltimore.

Calculations Net hepatic balance for plasma glucose, blood lactate, blood amino acids, and blood glycerol were determined by the formula ((0.28A + 0.72P)-H) x HF where A is the arterial concentration, P is the portal vein concentration, H is the hepatic vein concentration and HF is hepatic flow (blood or plasma) as determined by indocyanine green according to the method of Leevy.” Gut balance for these substrates was determined by the formula (A-P) x 0.72 HF. The fractional extraction of substrates by the liver was determined using the formula ((0.28A + 0.72P) -H)/(0.28A + 0.72P). In the case of glucose, the use of blood glucose and hepatic blood flow

OL-_I

6

TIME(h) Fig 1. Effects of consumption of a mixed meal on arterial plasma insulin and glucagon concentrations in 96hour fasted conscious dogs. Values are means + SEM for five dogs. lP < .05.

858

DAVIS, WILLIAMS, AND CHERRINGTON

MEAL

ARTERIAL

I50

GLUCOSE

100

10.0

cI

0"TP"T

_I_

GLUCOSE

(mg/kg.min)

- 1.0 -2.o-

I

-3.0-l

GUT LACTATE BALANCE

5.0 ’

40.0

4.0r I

LL

I

i

I

SPLANCHNIC GLUCOSE BALANCE (mg/kpmin)

-l.OL~ ’ -I 0

I

I

2 TIME

4 01)

I

i

6

6

-loot -15aL

Fig 2. Effects of consumption of a mixed meal on arterial plasma glucose concentration and net hapatic. gut, and splanchnic glucose balances in 96-hour fasted conscious dogs. Values ara means f SEM for five dogs. lP < .05.

y-I ’ -I 0 ’

I 2 TIME

I

1

1

4 Ihl

6

6

Fig 3. Effects of consumption of a mixed meal on the arterial blood lactate concentration, net hepatic, gut, and splanchnic lactate balances in 96-hour fasted conscious dogs. Values ara means + SEM for five dogs. lP < .05.

Lactate Metabolism Arterial blood lactate rose from a control value of 678 + 71 to 1,000 f 158 pmol/L (P -C .05) 30 min after feeding and continued to rise such that by 2 hours it was 1,83 1 + 115 pmol/L (Fig 3). It then declined but was still significantly elevated at 8 hours (1,279 + 72 pmol/L). Lactate output by the gut was not significantly altered by feeding. Hepatic lactate balance, on the other hand, changed from a net uptake of 12.19 + 2.02 pmol/kg/min to a net output of 4.81 f 1.71 hmol/kg/min (P -C .05) by two hours. Lactate output by the liver remained elevated for the next 6 h and averaged 5.21 * 0.05 pmol/kg/min.

Glycerol Metabolism Arterial blood glycerol decreased from a control period mean of 130 i 14 to 61 i 11 pmol/L (P -C .05) 30 minutes after feeding and then remained suppressed (Fig 4). There was no significant uptake or output of glycerol by the gut at any time during the experiment. The liver, however, consumed glycerol at a rate of 3.04 _t 0.26 pmol/kg/min during

the control period but at a rate of only 1.53 + 0.37 kg/min (P < .05) thereafter. The hepatic fractional tion of glycerol (Table 1) decreased slightly from mean of 0.67 + 0.03 to 0.53 k 0.05 (P < .05) minutes.

pmol/ extraccontrol by 30

Amino Acid Metabolism The arterial level of the gluconeogenic amino acids (alanine, glycine, serine, threonine, glutamine, and glutamate) increased from 1470 k 39 to 1912 + 95 pmol/L (P < .05) within one hour of feeding and remained elevated until the end of the experiment (Fig 5, Tables 1 and 2). The gut switched from a net uptake of the gluconeogenic amino acids (2.19 + 1.58 pmol/kg/min) to a maximal output of 11.62 +1.30 Fmol/kg/min, (P -C .05) four hours after feeding. The net uptake of the gluconeogenic amino acids by the liver, on the other hand, was elevated almost threefold one and two hours after the meal. It then fell to a rate somewhat less than twice the basal rate for the remainder of the study. The

NET HEPATIC LACTATE BALANCE

ARTERIAL

, 5O

GLYCEROL

,oo

BKIOD

(~molas/l

1

xl

t

ARTERIAL BLOOD GLUCONEOGENIC AMINO ACIDS

50 c

1

HEPAT IC GLUCONEOGENIC D AMI NO -5.0 AC I D BALANCE -10.0

GUT GLYCEROL BALANCE

-----J E OUTPUT

0

(rmolrdkg~mid

-25 -5.0

-4 i=

UPTAKE

L i

I

I

1

I

I

GUT GLUCDNEDMNIC AMINO ACIDBALANCE tjrmotewlrqmin)

SPLANCHNIC GLYCEROL BALANCE (pmolcr/kqvin)

15.0 IUD

50 c

I

I

I

1

2

4

6

8

Fig 4. Effects of consumption of a mixed meal on the arterial blood glycerol concentration, net hepatic, gut. and splanchnic glycerol balances in %-hour fasted conscious dogs. Values are means t SEH for five dogs. ‘P < 335.

fractional extraction of the gluconeogenic amino acids by the liver doubled after feeding (Table 1) although the change was not significant.

I

I

I

I

-5.0

*or

I 0

1

cT t

I -=.o _,

I

1OUTPUT

I UPTAKE

-lO.OL’ I 2.“F[7 _2.5

I

SPLANCHNIC “‘GLUCONEOGENIC u AMINO t ACID -50 BALANCE fpmolca/lq~minl -lOfl [WI _:

I

1 I

3

OUTPUT

I UPTAKE I 0

2

4 TIME thl

L 6

e

Fig 5. Effects of consumption of a mixed meal on the combined arterial gfuconeogenic amino acid (alanine, glycine, threoninc. serine, glutamate and glutamine) concentration, net hepatic. gut. and splanchnic gluconeogenic amino acid balances in 96-hour fasted conscious do9s. Values are means + SEM for five dogs. lP < .05.

Glutamine In the case of glutamine, the arterial level (Table 2) actually decreased after feeding (from 660 + 27 to 560 f 24 pmol/L, P < .05, by eight hours). Gut glutamine balance switched from a net uptake of 1.54 A 0.24 pmol/kg/min during the control period to a maximum output of 1.01 +0.49 pmol/kg/min 6 hours after feeding (P c .OS). The balance of glutamine across the liver, on the other hand, changed from a net output of 1.30 + 0.48 rmol/kg/min to an uptake of 2.36 r 0.73 gmol/kg/min (P c .05) 1 hour after the meal. Branched Chain Amino Acids The arterial increased from pmol/L by 4 elevated at 8

blood level of the branched chain amino acids 43 I + 28 pmol/L to a maximum of 660 k 36 hours (P < .05) after feeding and was still haurs (Table 3). Net gut balance of the

branched chain amino acids changed from an uptake of 0.35 f 0.22 pmol/kg/min (NS) to a peak output of 3.12 2 0.70 rmol/kg/min (P < .05). Net branched chain amino acid balance across the liver also changed from an uptake of 0.35 k 0.58 wmol/kg/min (NS) to a peak output of 2.32 + 1.38 qnoi/kg/min fP c .05) during the 8 hours after feeding. Blood Urea Nitrogen Metabolism The arterial blood urea nitrogen (BUN) level increased from 2,370 i 280 pmol/L to a maximum of 4,180 e 290 pmol/L five hours after the meal and remained elevated until the end of the experiment. There were no significant changes in gut BUN balance, but the hepatic BUN output had increased fourfold two to three hours after the meal (Table 4).

860

DAVIS, WILLIAMS,

Table 1. Effect of a Mixed Meal*

on Hepatic Fractional

of Blood Glycerol and the Blood Gluconeogenic in 96-Hour

Table 3. Effects of a Mixed Meal* Splanchnic

2

4

Fasted Conscious Dogs (n = 5)

8

Control Period

x

0.66

0.53t

0.54t

0.50t

0.57t

2 SEM

0.03

0.04

0.03

0.03

0.02

Arterial concentration (pmol/L) Hepatic balance

Gluconeogenic

x

0.10

0.17

0.21

0.10

0.10

amino acids

z~ SEM

0.03

0.04

0.07

0.06

0.05

Cpmol/kg/min)

*Meal fed at 0 time.

j;

431

+ SEM

28

X -0.35 ? SEM

Gut balance (rmol/

tP < 0.05.

kg/min)

+ SEM

(pmol/kg/min)

0.22

j; -0.70 t SEM

0.47

Hours 1

2

4

581’

657.

660.

39

8 608*

32

36

26

2.32t

2.10t

1.31

1.07

1.38

0.66

0.53

1.20t

2.53t

3.12t

2.94t

0.65

1.07

0.70

0.54

3.01t

4.86t

5.26t

4.26t

0.65

1.30

0.44

0.73

1.82t

Negative sign indicates net uptake.

The total hepatic glycogen content of dogs which were fasted 96 hours and then killed rather than studied was 6.3 + 1.7 g. Dogs of similar size and nutritional status, which were fasted 96 hours, fed a mixed meal, and killed 8 hours later, had a hepatic glycogen content of 3 1.5 + 1.2 g. Thus, in the first 8 hours after a mixed meal was fed to the 96-hour fasted dog, approximately 25 g of hepatic glycogen were synthesized. Previous experiments indicated that during the first 12 hours after ingestion of a similar meal by overnight fasted dogs 44 g of glycogen were synthesized, a change from 35 f 4 g to 79 Z!Z15 g.” DISCUSSION

In contrast to the overnight fasted dog,’ 96-hour fasted dogs exhibited marked net hepatic uptake of lactate during the control period of the study. In association with this they had significantly less hepatic glycogen and a lower overall glucose production rate. Clearly after a 96-hour fast in the dog the net movement of lactate is from the periphery to the liver rather than the reverse as is the case in the overnight fasted dog.’ These data are in line with our earlier finding that the healthy conscious dog does not begin to exhibit net hepatic lactate uptake until 22 to 24 hours after feeding.’ Most importantly the data from the present experiments indicate that within 90 minutes of mixed meal feeding the liver of the 96-hour fasted dog switches from net lactate consumption to net lactate production. The lactate response to the meal was similar both in magnitude (ie, the change

Table 2. Effects of a Mixed Meal.

on the Arterial

of Glutamine

*Meal fed at 0 time. tP < .05.

from baseline) and time of onset (ie, minutes after feeding) to that observed previously in the overnight fasted dog.’ Clearly, therefore. in the four-day fasted dog, as in the overnight fasted dog, lactate, in a net sense, cannot be a contributor to hepatic postprandial glycogen synthesis. The source of the carbon for the lactate produced and the glycogen synthesized in the postprandial period remains unclear. It seems unlikely that circulating glucose contributed carbon in a net sense, since although hepatic glucose production was suppressed, the liver did not exhibit net glucose uptake, confirming a similar finding in the overnight fasted dog fed a mixed meal’ or a meat meal.14 This lack of net glucose uptake can perhaps best be explained by the failure of the plasma glucose level and therefore the hepatic glucose load to reach a critical value.” In addition the rise in glucagon may have countered the effects of insulin and glucose on net hepatic glucose uptake.‘* It is possible that our measured portal glucose concentrations underestimated the true portal glucose levels as a result of poor mixing of the ingested glucose in the laminar flow of the portal vein. Although this would have caused an underestimate of net hepatic glucose uptake it seems unlikely for several reasons. First, any mixing errors should have been random and thus should not have been apparent in the mean data. Second, in a previous study in which dogs with cathe-

Blood Glutamine

in 96-Hour

Level and Net Hepatic. Gut, and Splanchnic

Arterial concentration

x

Balances

Fasted Conscious Dogs (n = 51 HOWS

Control

(~mol/L)

0.58

x -0.35

Splanchnic balance

Hepatic Glycogen Content

Blood Branched

and Net Hepatic. Gut, and

Balances of the Branched Chain Amino Acids

in 96-Hour

HoursAfter Feeding 1

on the Arterial

Chain Amino Acid Concentration

Fasted Conscious Dogs (n = 5) Control Period

Glycerol

Extraction

Amino Acids

AND CHERRINGTON

Period

1

2

660

618

586

27

x

1.30

(~mollkglmin)

f SEM

0.48

0.73

1.72

0.62

1.10

Gut balance (pmoll kgjmin)

x + SEM

-1.54 0.24

-0.74t

0.50t

0.64t

1.01t

0.24t

0.45

0.17

0.49

0.44

-0.24

-3.10

(flmol/kg/min)

ii + SEM

Negative sign indicates net uptake. *Meal fed at 0 time. tP < .05.

0.4 1

0.22 0.91

37

33 - 1.38*

-1.46 0.34

-0.74 1.06

49

8 560t

f SEM

-2.36t

- 1.96t

6 5151

Hepatic balance

Splanchnic balance

30

4 544’

-0.63t

0.38 0.59

24 -0.87t 0.70

-0.63 0.72

861

NET HEPATIC LACTATE BALANCE

Table 4.

Effects of a Mixed Meal.

on the Arterial

Blood Urea Nitrogen

of Blood Urea Nitrogen

in g&Hour

1.5

Arterial concentration

x

2.370

3,000

Concentration

and net Hepatic, Gut. and Splanchnic

2.5

3.5

3,760t

3,890t

5 4,180t

(Irmol/L) Hepatic balance

+ SEM

280

310

260

270

320

ii

3.70

12.40t

14.70t

10.70t

12.70t

(flmollkglmin)

_t SEM

0.40

x

Gut balance (amollkgl min)

-3.00

4.80

4.30

- 2.00

- 3.60

4.30 -2.30

f SEM

0.56

0.90

0.80

31.0

2.50

x

2.70

9.40t

12.70t

7.10t

10.40t

f SEM

0.66

3.10

2.30

3.60

2.20

Splanchnic balance (pmol/kg/min)

1.90

-0.96

Balances

Fasted Conscious Dogs ht = 5)

7 4,lBOt 290 3.10 3.30 0.32 1.BO 3.42 2.00

Negative sign indicates net uptake. *Meal fed at 0 time. tP < .05.

ters placed in a similar manner were used, 96% of an oral glucose load was recovered over the three-hour absorptive period. I9 Possible errors relating to catheter placement or blood flow measurements (note that liver blood did not change significantly, Table 5) in the calculation of net gut or hepatic balance of glucose or other metabolites have been discussed previously.’ Only a third of the digestable glucose administered in the meal can be accounted for, but this failure to absorb the load completely relates to the fact that absorption was not complete in eight hours. This is clear from the observation that even eight hours after feeding the rate of glucose output was not significantly different from the peak rate evident at six hours. In addition a significant portion of this glucose load was probably metabolized by the gut itself.” One carbon source undoubtedly used for glycogen synthesis and lactate production by the liver is that of the gluconeogenic precursors. The present experiments indicate that net hepatic uptake of the gluconeogenic amino acids (alanine, glycine, serine, threonine, glutamate, and glutamine) and glycerol can account for as much as of 25% of the required carbon. The hepatic uptake of small peptides derived from the diet would provide another source of carbon. Absorption of small peptides has been shown to occur after a horsemeat meal,20 and if some absorption of peptides did indeed occur, it is possible that their carbon skeletons could have been used by the liver for glycogen repletion. It is also possible that carbon could have been derived from nonglucose carbohydrates in the meal, since only half of the total reducing sugars found in the diet after acid hydrolysis could be accounted for by glucose. The other remaining possibility is an intrahepatic carbon source. If such were the case, it is unlikely to be stored lipid as net glucose synthesis from fatty acids does not occur and Table 5. Hepatic Blood Flow in S&Hour

Fasted Conscious Dogs

Fed a Mixed Meal at 0 min (n = 51 Minutes -30

-15

0

15

60

120

180

240

360

480

Hepatic

blood flow (mL/kg/min)

38

39

39

42

38

38

39

42

41

41

3

3

3

4

3

3

2

2

2

2

the liver most likely does not contain sufficient lipids for the hydrolyzed glycerol to make a substantial contribution. This leaves open the possibility that hepatic proteolysis provided some of the carbon for glycogen synthesis. It has been shown that glucagon, which increased rapidly in these experiments, can stimulate proteolysis in the liver.2’ Also consistent with enhanced proteolysis following the meal is the fact that there was a significant postprandial release of branched chain amino acids, usually an indication of proteolysis. Furthermore, the amount of urea which was produced by the liver after the meal was more than 50% greater than could be accounted for by the hepatic uptake of amino acids. On the other hand work by Mortimore et a122indicated that elevation of the plasma amino acid levels can inhibit hepatic proteolysis. In addition high levels of insulin might be expected to slow hepatic proteolysis. Unfortunately, the present study does not allow resolution of these various effects and does not provide direct data regarding the issue of postprandial hepatic proteolysis. In these experiments glutamine was measured along with the other individual amino acids. The effect of fasting on glutamine metabolism is reflected in a slight increase in the arterial blood glutamine level. This increase is probably attributable to the fact that net hepatic glutamine balance changed from a slight uptake in the overnight fasted state to output after a longer fast.2’ Such a change might occur if, as a fast progresses, glutamine becomes less important as a hepatic gluconeogenic precursor and more important as a vehicle for transporting amino groups, which were derived from the deamination of gluconeogenic amino acids, to the kidney. Since the rise in the arterial glutamine level is so small, it suggests that its uptake by the periphery, the kidneys in particular, increases in parallel with the elevated hepatic glutamine production. This is consistent with the finding by Cersosimo et a123that between 24 and 96 hours of fasting the renal bed switches from a slight producer of glutamine to a net consumer. However, as these data show, upon refeeding, the liver begins to take up glutamine, rather than produce it, resulting in a decline in the arterial level of the amino acid. Thus, following a meal, even after a 96-hour fast, glutamine apparently again assumes a more prominent role as a gluconeogenic precursor. It is of interest to note that glutamine infusions in the

862

DAVIS, WILLIAMS,

96-hour fasted conscious dog also resulted in the liver becoming a net consumer of glutamine rather than a producThis finding is consistent with our data in that an er. 23S24 increase in the concentration of glutamine entering the liver resulted in hepatic uptake of the amino acid. In summary, the data from these experiments indicate that in a conscious, 96-hour fasted dog: (1) the liver begins to produce lactate following consumption of a mixed meal, (2) net hepatic glutamine balance changes from production to

AND CHERRINGTON

uptake following consumption of a mixed meal, and (3) net hepatic glucose production is reduced but net hepatic glucose uptake does not appear to occur. This study further brings into question the source of carbon for postprandial glycogen synthesis. In that regard, however, it must be remembered that our goal was to study net hepatic lactate balance not the carbon source for liver glycogen; thus, we did not administer a meal of defined composition and our data regarding the carbon source for glycogen synthesis await confirmation.

REFERENCES

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