Effect of somatostatin and a glucagon-specific analog on glucose homeostasis during arginine infusion

Effect of Somatostatin and a Glucagon-Specific Analog on Glucose Homeostasis During Arginine Infusion Eric L Lien The glucose

response

normal

was

rats

glucagon 1

to arginine

studied

deficiency

mg/kg/hr)

ciency

insulin

(somatostatin

or

selective

In control

glucagon

cose

rose

levels

arginine

14

and returned

termination

of

the

increased

136

increased

76 *

Infusion

*

12

pg/ml

administration during associated

glu-

modest

Insulin

levels

demonstrate

during

somatostatin

the

do

infu-

resulted

rise in insulin that

not

arise

cagon-specific

in

produced It

glucagon

of somatostatin

glucagon

and The

[ D-Cys”]somatostatin infusion

of

insulin

release,

ensued.

hyperglycemia.

suppression

to

glucagon

of

arginine

at the

and

arginine-induced

hyperglycemia

levels

to basal KU/ml

marked 1

in response

infusion.

12

of

mg/dl

plasma

of both

and arginine-induced

defi-

infusion,

studies,

supression

in and

infusion,

([D-Cys”]somatostain

mg/kg/hr).

sion.

infusion

during

secretion

release.

These

treated

a

results

the hyperglycemic

somatostatin

in

and effects

in arginine-treated in animals

no

resulted

animals with

glu-

analogs.

T

HE CYCLIC tetradecapeptide somatostatin inhibits the release of numerous hormones, including those from the pituitary (growth hormone’ and prolactin’), the pancreas (insulin” and glucagonl) and the gut (gastrin,’ cholecystokinin.” and secretin’). One potential therapeutic use of somatostatin would be to lower inappropriately high glucagon levels in the diabetic state which may contribute to the hyperglycemia of this disease.’ Studies with insulin dependent diabetics have indicated the ability of somatostatin infusions to lower postabsorppatients. Intive blood glucose levels!‘,“‘and prevent ketosis I1 in insulin-withdrawn fusion of somatostatin in maturity-onset diabetics and normal subjects results in most likely due to suppression of inmodestly elevated blood glucose levels,“-“’ sulin release. The use of glucagon-specific analogs, such as those developed by Brown et al.‘; ([D-Cys”]-somatostatin and [D-Trp’, D-Cys”]-somatostatin), may prevent elevation of blood glucose levels observed after somatostatin infusion. The present studies were undertaken to examine the effects of somatostatin and [D-Cys”]-somatostatin on insulin, glucagon and glucose levels during arginine in normal rats. MATERIALS Male Charles In order

to maintain

was cannulated matostatin @nine

(I I

anesthesia

(I

hourly

AND

injections

of Nembutal.

and rats were treated

mg/kg/hr,

from

six animals)

from

30 60 min. or (3) saline infusion

animals).

Blood samples

and

Metabol/sm,

1.2mg

75.90.

were obtained

and I20 min after

of versene/0.5

Vol 27. No 9 &?ptember).

METHODS

300~ 350 g were anesthetized

30 60 min, (2) [D-Cys”]somatostatin

30 set) 0, 15. 30,45,60, Trasylol

CD rats weighing

for infusions

infusion

7.2) from min

River

with Nembutal,

IO mg/kg.

with one of the following six animals)

from 0 30 min followed from

three

time 0 60 min plus arginine

(I mg/kg/hr. the jugular

(i.p.).

by arginine

vetn

(I) so-

protocols:

(100 mg/kg/hr.

from

vein (a procedure

the start of the infusion

50 mg/kg

were given. The jugular

pH

time 0 60 min plus alone from requiring

in Trasylol-versene

30 60

less than (600 U of

ml of blood).

1978

1095

1096

ERIC L. LIEN

Hormone levels were measured by radioimmunoassay. Insulin was determined by the method of Hales and Randle’” using human insulin standards (Schwarz/Mann) and glucagon was determined by the method of Faloona and Unger19 utilizing Unger 30K glucagon antiserum. Glucose was determined by the glucose-oxidase method. Somatostatin was synthesized by solid-phase methodology:“” [D-Cys”]-somatostatin was a gift of Dr. J. Rivier. RESULTS

During an arginine infusion in control animals plasma glucose increased 14 mg/dl from a mean (‘t SEM) of 79 =t 3 mg/dl during saline infusion to 93 f 4 mg/dl (Fig. 1). After termination of the arginine infusion, glucose returned to basal levels. An unexplained rise of glucose levels to 92 f 3 mg/dl was observed at the termination of the experiment. The combination of somatostatin and arginine infusions resulted in marked hyperglycemia; glucose levels peaked after the cessation of arginine infusion (159 f 17 mg/dl) and did not return to normal until 60 min later. Glucose levels during [D-C~S’~]-somatostatin infusion were similar to control values (Fig. 1) during basal and combined arginine and [D-Cys”]somatostatin administration. Glucose levels did not, however, return to preinfusion levels following termination of the infusion, but instead slowly rose to 113 i 3 mg/dl at the end of the experiment. Plasma insulin levels in control animals responded rapidly during the arginine infusion, increasing 136 i 12 /*U/ml after 15 min of arginine administration (Fig. 2); following the arginine infusion insulin rapidly returned to basal levels. Infusion of somatostatin resulted in a rapid initial drop in insulin levels, a slow rise during the arginine infusion and a marked rebound after the termination of the somatostatin-arginine infusion. [D-Cys”‘]-somatostatin administration resulted in a reduction of basal insulin levels similar to that seen with somatostatin. During the arginine-[D-Cys”]-somatostatin infusion insulin levels rose, but not to the same SRIF

or

D-c~s’~-SRIF

ARGININE

170

l

* I

\

160

150

140

\\ \\ \\ \\

I’ I’

0

30

60 RIINUTES

90

\ 120

Fig. 1. Effect of somatostatin (1 mg/kg/hr) or [D-Cys’4]-somatostatin (1 mg/kg/hr) infusion on plasma glucose during arginine infusion (100 mg/kg/hr). D--O: control (n = 111; e---e: somatostatin (n = 6); x.. .x: [D-Cys”]-somatostatin In = 6). Vertical bars represent SEM; p < 0.01, + p < 0.06. The p values refer to peptide treated animals when compared to saline controls.

GLUCOSE

HOMEOSTASIS

DURING

ARGININE

INFUSION

1097

SRIF or D-ryd4-SRIF ,

ARGININE

,

160,-

Fig.

2.

Effect

mg/kg/hr) (1

or

mg/kg/hr)

sulin

somatostatin on

arginine o---i>:

plasma

infusion control

somatostatin

0.01,

bars + p <

to peptide-treated pared to saline

The

SEM; p values

animals

11);

.x:

x.. (n

represent 0.05.

in(100

(n =

(n = 6):

[ D-Cys14]-somatostatin Vertical

(1

[D-Cys”]-somatostatin infusion

during

mg/kg/hr). e----e:

of

I

when

=

61.

p < refer com-

controls.

extent as when arginine was infused alone. The insulin levels in animals receiving arginine and [D-Cys’)]-somatostatin were significantly higher than somatostatinand arginine-treated animals (p < 0.01 at 45 and 60 min), and significantly lower than animals receiving arginine alone (p < 0.01 at 45 and 60 min). Plasma glucagon levels rose 76 f 16 pg/ml in control animals in response to arginine. The gfucagon levels in animals treated with either somatostatin or [D-Cys’l]-somatostatin were lowered during the basal period (since control levels also declined during this time, no significant differences were noted) and did not increase in response to arginine (Fig. 3). 100

SRIF or D-cyri4SRIF 1

ARCININE

,

Fig.

3.

Effect

mg/kg/hr)

or

(1 mg/kg/hr) cagon mg/kg/hr.

somatostatin

infusion

during

e----e:

of

on

arginine

~i--:~:

control

somatostatin

Vertical

bars + p <

to peptide-treated pared

to saline

= 11); x..

In The

SEM: p

animals

controls.

glu(100

(n

(n = 6);

represent 0.05.

plasma

infusion

[D-Cys”]-somatostatin 0.01,

(1

[II-Cys”]-somatostatin

.x:

= 6). p <

values refer when

com-

1098

ERIC L. LIEN

DISCUSSION

The present study was undertaken to determine the effects of suppression of glucagon and insulin release (by somatostatin) and the selective suppression of glucagon (by [D-Cys”]-somatostatin) on the arginine-induced elevation of blood glucose. Previous studies have demonstrated the diabetogenic action of somatostatin during arginine tolerance tests in normal humans and maturity-onset diabetics.12 The same type of prolonged hyperglycemia was found when somatostatin and arginine were infused simultaneously in rats (Fig. 1). The arginine-stimulated rise of insulin and glucagon was blocked by somatostatin, as seen in Figs, 2 and 3: however, the decrease in insulin was mare pronounced than in glucagon. Hyperglycemia was probably a result of the relative excess of glucagon, capable of stimulating both glycogenolysis” and gluconeogenesis,” and a reduced rate of glucose clearance due to lowered insulin levels. Somatostatin and [D-Cys”]-somatostatin differ in their ability to suppress pancreatic hormone secretion; Brown et al. found the analog to be more potent than somatostatin in inhibiting glucagon release, while it only weakly inhibited inobvious in this study: alsulin release.” This relative specificity was immediately though the arginine-induced glucagon release was completely inhibited, insulin levels were intermediate between those of controls and somatostatin-treated animals. Following the infusion, glucagon levels rebounded while insulin levels rose modestly (not as severe a rebound as with somatostatin), and a mild elevation of glucose levels ensued. The ability of somatostatin infusion to lower blood glucose!‘.‘” and prevent ketoacidosis” in insulin-withdrawn juvenile diabetics has previously been demonstrated. Less desirable results were obtained when somatostatin was infused in normals or in maturity-onset diabetics. Prolonged somatostatin infusion in normal postabsorptive humans’“,‘+ and maturity-onset diabetics’” resulted in a transient fall in glucose levels followed by a sustained rise to levels significantly greater than the basal values. Somatostatin infusion also decreased the glucose utilization rate following intravenous glucose tolerance tests in humans.“?,‘” Arginine tolerance tests in normal humans during somatostatin infusions have produced several responses. Gerich et al.4 found no increase in glucose levels during infusions of arginine plus somatostatin, but the infusions were followed by hyperglycemia. The dose of somatostatin employed was sufficient to suppress arginine-induced insulin and glucagon release (glucagon levels actually decreased transiently during the infusion); a glucagon rebound followed the termination of the somatostatin infusion. Changes observed in the glucose levels were probably mediated by changes in the glucagon levels: both parameters were suppressed during the arginine-somatostatin infusion, and rebounded following the infusion (insulin remained unchanged during the experiment). WaldhPusl et al.,” employing a higher dose of arginine and a lower dose of somatostatin, found concomitant administration of arginine and somatostatin resulted in prolonged hyperglycemia when compared to the administration of arginine alone. Somatostatin infusion completely suppressed arginine-stimulated insulin release, while glucagon rose slightly. The relative excess of giucagon, stimulating hepatic glucose output, coupled with lowered glucose clearance due to suppressed insulin levels probably resulted in the observed hyperglycemia (results similar to those in the present study). Administration of arginine and [D-Cys’4J-somatostatin in the present

GLUCOSE

HOMEOSTASIS

DURING

ARGININE

INFUSION

1099

study did not result in elevated glucose levels, although the insulin response was attenuated. The absence of hyperglycemia in this experiment is therefore most likely due to complete suppression of the glucagon response. Since glucose and pancreatic hormone levels react similarly to combined somatostatin and arginine infusions in rats (present experiment) and in normal humans and maturity-onset diabetics,” the result with glucagon specific somatostatin analogs should apply to humans. Glucagon levels of maturity-onset diabetics are inappropriately elevated in response to a number of stimuli. Although a large carbohydrate load reduced glucagon levels in normals, it failed to do so in maturity-onset diabetics despite marked hyperglycemia.‘” Prolonged arginine infusion resulted in excessive glucagon levels in maturity-onset diabetics when compared to contro1s,L4.‘)” and insulin failed to lower arginine-stimulated glucagon levels.“’ Although [D-Cys”]somatostatin does inhibit basal insulin release in addition to its potent suppression of arginine-stimulated glucagon release, an analog with even greater glucagon specificity is expected to selectively reduce glucagon levels in maturity-onset diabetics in both the basal and stimulated states, and thus improve glucose homeostasis. ACKNOWLEDGMENT I wish to thank Gerald

Deitch and Ruth Gillis for their excellent

technical

assistance

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ERIC L. LIEN

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