Physiological responses to acute stress in alloxan and streptozotocin diabetic rats

Physiology & Behavior, Vol. 45, pp. 483-489. ©Pergamon Press plc, 1989. Printed in the U.S.A.

0031-9384/89 $3.00 + .00

Physiological Responses to Acute Stress in Alloxan and Streptozotocin Diabetic Rats JANA H. LEE, MARIA

KONARSKA

AND RICHARD

McCARTY 1

D e p a r t m e n t o f P s y c h o l o g y , University o f Virginia, Charlottesville, VA 22903 R e c e i v e d 22 S e p t e m b e r 1988 LEE, J. H., M. KONARSKA AND R. McCARTY. Physiological responses to acute stress in alloxan and streptozotocin diabetic rats. PHYSIOL BEHAV 45(3) 483-489, 1989.- Physiological responses to acute stress were assessed in alloxan diabetic, streptozotocin diabetic and control laboratory rats. Rats were prepared with indwelling tail artery catheters to allow for direct measurement of mean arterial pressure (MAP, mmHg) and heart rate (HR, beats per minute) and remote sampling of blood. Within 24 hours after surgery, basal values of MAP and HR were determined. Two days after surgery, rats were subjected to 5 minutes of intermittent footshock. Blood samples were collected before footshock stress and immediately and 15 minutes after termination of footshock. Plasma levels of norepinephrine (NE) and epinephrine (EPI) increased significantly above basal values in all animals exposed to acute footshock stress. However, in approximately one-half of alloxan and streptozotocin diabetic rats, plasma levels of EPI under basal conditions and following footshock stress were elevated significantly compared to controls and the remaining diabetic animals. We have denoted these subgroups of diabetic animals as reactive responders (plasma EPI greater than controls) and nonreactive responders (plasma EPI similar to controls), respectively. Plasma levels of NE under basal conditions and following footshock stress were similar in reactive responders and nonreactive responders compared to matched controls. Baseline blood glucose levels were elevated in alloxan and streptozotocin diabetic rats compared to controls. Blood glucose levels increased reliably in all animals following footshock stress. Basal MAPs were reduced significantly in both subgroups of alloxan and streptozotocin diabetic rats compared to matched controls. In contrast, resting HRs were similar between diabetic rats and their corresponding controls. These data indicate that a distinct subgroup of animals with chemicallyinduced diabetes exists which maintains elevated basal circulating levels of EPI and has exaggerated plasma EPI responses to acute stress. These reactive responders may have greater difficulty with glycemic control, especially during periods of stressful stimulation. Plasma catecholamines Animal models

Epinephrine

Norepinephrine

C H R O N I C elevations of blood glucose are the most prominent feature of the diseases termed diabetes mellitus. Diabetic complications are a leading cause of blindness, kidney failure, congenital m a l f o r m a t i o n s and amputations o f the lower extremities. Diabetes also constitutes a m a j o r risk factor for cardiovascular disease (8). It is widely thought that poor glycemic control exacerbates these complications. Hence, much attention has been focused upon the effects of stressful stimuli on regulation o f blood glucose levels and overall diabetic control. In human diabetics, stress effects on diabetic control may alter glucoregulatory processes or disrupt adherence to therapeutic regimens (9,19). It is generally assumed that acute exposure to stressful stimulation is attended by increases in blood glucose secondary to counterregulatory h o r m o n e release. Thus, blood glucose responses to acute stress would be expected to be greater in subjects with insulin dependent diabetes mellitus (IDDM) since little or no endogenous insulin is secreted to offset acute increases in blood glucose (7). In the clinical literature, there is a lack of evidence to suggest that exposure to stress results in consistent increases in

Alloxan

Streptozotocin

Diabetes

Stress

blood glucose levels (3, 4, 7, 12). For example, a review by Lustman and co-workers (21) indicates that certain experimental stressors such as electric shock are associated with acute hypoglycemia. More recently, Kemmer and co-workers (14) reported that the stress of public speaking and mental arithmetic produced significant elevations in blood pressure, heart rate and plasma catecholamine concentrations but no changes in blood glucose. Systematic research on acute stress in animal models of Type I diabetes (insulin dependent) is even more limited. In one study, control and alloxan diabetic mice were exposed to groups of aggressive conspecifics. In control animals only, blood glucose levels increased significantly following the stressful stimulation (2). In another study, Carter and co-workers found that baseline blood glucose levels served as a predictor of blood glucose changes following rotation stress in alloxan diabetic animals. That is, when animals were tested when hyperglycemic, blood glucose levels decreased following rotation stress and vice versa. In addition, blood glucose levels of control animals did not change significantly following rotation stress (4). In other studies, blood glucose levels of control ani-

1Requests for reprints should be addressed to Richard McCarty, Ph.D., Department of Psychology, 102 Gilmer Hall, University of Virginia, Charlottesville, VA 22903-2477.

483

484

LEE, KONARSKA AND McCARTY

mals have been reported to increase following stressful stimulation (35). Several methodological problems limit firm conclusions regarding the role of stress in IDDM and in glucoregulation. These include: (a) inappropriate timing of blood samples in relation to stressful stimulation; (b) variability in the use of animal models of diabetes; (c) use of limited physiological measures of stress responses and (d) lack of consistency in the use of stressful stimuli. In the present study, we have examined several physiological responses to acute footshock stress in control, alloxan diabetic and streptozotocin diabetic rats. We were especially interested in comparing plasma catecholamine and blood glucose responses to acute stress in two animal models of diabetes. EXPERIMENT 1 METHOD

Animals Adult male Sprague-Dawley rats (325-350 g) were purchased from Hilltop Breeding Laboratories, Scottdale, PA. Rats were housed individually in Wahmann suspended metal cages with laboratory chow and tap water freely available. Animals were allowed to acclimate to the vivarium for at least 1 week prior to testing. The vivarium was maintained automatically on a 12 hour light-dark cycle (lights on at 0700 hours) at a temperature of 22°C.

Alloxan Treatment Following a 48 hour fast, 10 rats received intraperitoneal (IP) injections of alloxan monohydrate (120 mg/kg) dissolved in 0.9% sodium chloride. Fourteen control animals received IP injections of 0.9% sodium chloride (2.5 ml/kg). Within 72 hours following alloxan administration, blood glucose concentrations were measured via tail clip sampling and an Accu-Chek II reflectance meter (Boehringer Manneheim Diagnostics, Indianapolis, IN). Blood glucose levels were greater than 250 mg/dl in all alloxan-treated animals. Beginning on the third day after alloxan treatment, diabetic rats received daily injections of regular insulin (2 U/animal, SC) between 1700-1900 hours.

Tail Artery Catheters Within 12 days after alloxan or vehicle treatment, animals were anesthetized with methylhexitol (Brevital, 45 mg/kg, IP) and a PE50 catheter was surgically placed into the ventral tail artery according to the method of Chiueh and Kopin (6). Briefly, an incision was made into the tail sheath, the ventral caudal artery was dissected free and elevated, and a PE50 catheter was inserted up into the artery and secured with two size 00 sutures. The tubing was run subcutaneously to exit in the midscapula region and was protected by a stainless steel spring wire and adhesive tape collar. The catheter was filled with heparinized saline (300 I.U. per ml) and the end of the tubing was occluded with a 23 G needle and disposable tuberculin syringe. After recovery from surgery, animals were housed individually in plastic cages (35 x35 x22 cm) that contained a layer of bedding material and ad lib supplies of food and water. The catheter and spring wire were led out of the top-center of the cage and secured such that each rat could move freely throughout the cage. Catheters were flushed in the early morning and late afternoon with 0.5 ml of heparinized saline to maintain patency. On the day after surgery, direct measures of mean arterial

pressure (MAP, mmHg) and heart rate (HR, beats per minute) were obtained by connecting the end of the catheter to a Statham pressure transducer, with tracings made on a Grass multichannel polygraph. HR was measured by an arterial pulse triggered cardiotachometer.

Acute Stress Testing Two days after surgery, diabetic rats received regular insulin (2 U per animal, IP) at 0800 hours. Controls received injections of 0.9O7o saline. Basal blood samples (0.5 ml) were then collected from each animal beginning at 1200 hours under quiet conditions while animals were resting in their home cages. The volume of blood removed for this and each subsequent blood sample was replaced immediately by a slow infusion of heparinized saline (100 I.U. per ml). After basal blood samples were collected from all animals, rats were transferred individually to a Plexiglas shock chamber with a floor of stainless steel rods spaced 1 cm apart (23). Scrambled footshocks were delivered automatically (0.75 mA, 0.6 seconds duration every 10 seconds for 5 minutes) by a Coulbourn shock generator and constant current unit. Additional blood samples were collected immediately and 15 minutes after the termination of footshock. Blood glucose levels were determined as described above as blood samples were collected.

Assay of Plasma Catecholamines Blood samples were collected into iced 10x75 mm glass culture tubes and centrifuged at 4000 × g for 10 minutes at 4°C in a Sorvall refrigerated centrifuge. Aliquots of plasma were removed and stored at - 7 0 ° C until assayed for content of NE and EPI within 2 weeks by a radioenzymatic-thin layer chromatographic procedure (10,29).

Data Reduction and Analysis Results for all groups are expressed as means+_SEM. Levels of significance for body weights, cardiovascular measures, plasma catecholamines and blood glucose were determined by one-way ANOVAs. Post hoc comparisons between groups were made using Tukey-Kramer tests (15). Comparisons within animals were made using paired t-tests. RESULTS

Through exploratory data analysis, it was apparent that alloxan diabetic rats sorted into 2 distinct subgroups based upon their plasma EPI levels under basal conditions and following footshock stress. Those animals with elevated plasma EPI levels are termed reactive responders while those with plasma EPI levels that are similar to controls are termed nonreactive responders. All subsequent analyses employ these 2 subgroups of alloxan diabetic animals. Table 1 presents body weights of rats prior to alloxan or vehicle administration and at the time of surgery. Alloxan reactive and alloxan nonreactive animals had significant decreases in body weight over the time between drug administration and surgery (ps<0.05). In contrast, control animals maintained relatively constant body weights over the same period of time. Resting MAPs were significantly different among treatment groups, F(2,20)=5.42, p<0.05, with values for reactive and nonreactive diabetic animals significantly lower than values for controls (ps<0.05). There were no differences among groups in basal HRs (Table 2). Plasma levels of NE were similar in diabetic and control rats under basal conditions. Immediately after 5 minutes of foot-

A C U T E STRESS IN D I A B E T I C R A T S

485 1200 •

TABLE 1 BODY WEIGHTS (g) OF ALLOXAN-REACTIVE (ALX-R), ALLOXANNONREACTIVE (ALX-NR) AND CONTROL RATS A T THE TIME OF A L L O X A N OR VEHICLE ADMINISTRATION AND PRIOR TO SURGICAL PLACEMENT OF TAIL ARTERY CATHETERS

Group (N)

Pretreatment 404 + 17 439 ± 23 383 + 20

T 0L'J

800

•

REACTIVE NONREACTIVE

_z_~, °°°.

Presurgery 406 + 18 409 _+ 23* 360 ± 23*

•

E:

Z~ <~

Control (14) ALX-R (5) ALX-NR (5)

1000

O-CONTROL I-ALLOXAN~-ALLOXAN-

G_

400 200

•

0

BASAL

Values are means ± SEM for the indicated numbers of animals. *p<0.05 (paired t-test).

TABLE 2 BASAL VALUES OF MEAN ARTERIAL PRESSURE (MAP, mmHg) AND H E A R T RATE (HR, BEATS PER MINUTE) OF ALLOXAN REACTIVE (ALX-R), ALLOXAN-NONREACTIVE (ALX-NR) AND CONTROL RATS 1 DAY AFTER SURGICAL PLACEMENT OF TAIL ARTERY CATHETERS

POST FS

+ 1 5 MIN

SAMPLING TIME

FIG. 1. Plasma levels of norepinephrine in control, alloxan-reactive, and alloxan-nonreactive rats under basal conditions and immediately and 15 minutes after exposure to 5 minutes of intermittent footsbock. Values are presented as means and standard errors.

1200

EZ] - CONTROL -ALLOXAN- REACTIVE -ALLOXAN- NONREACTIVE

1000

Group (N)

MAP

HR 80(3

Control (14) ALX-R (5) ALX-NR (5)

109 _+ 2 94 + 6 98 _+ 4 p<0.05

333 ~: 6 341 ± 8 314 ___8 p>0.10

600

400

200

Values are means + SEM for the indicated numbers of animals. Levels of significance were determined by one-way ANOVAs.

0

BASAL

POST FS +15 MIN SAMPLING TIME

shock, plasma NE was elevated to a similar extent in rats of the 3 groups. At 15 minutes postfootshock, plasma NE was significantly different a m o n g groups, F(2,20)=3.50, p < 0 . 0 5 , with levels in reactive diabetic animals significantly higher than levels in nonreactive diabetes ( p < 0 . 0 5 ) (Fig. 1). There were significant intergroup differences in basal plasma levels of EPI, F(2,21)=4.90, p < 0 . 0 5 , with diabetic reactive responders having higher values than controls (p<0.05). Immediately following footshock stress, plasma EPI was significantly different among groups F(2,21)=18.33, p < 0 . 0 1 , with alloxan reactive animals having much higher plasma E P I levels than alloxan nonreactive or control rats (ps<0.05). There were also significant intergroup differences in plasma EPI at 15 minutes postfootshock, F(2,21)=8.05, p < 0 . 0 1 . Again, plasma EPI was much higher in alloxan reactive responders compared to animals o f the other two groups ( p s < 0 . 0 5 ) (Fig. 2). As summarized in Fig. 3, there were significant group differences in blood glucose under resting conditions, F(2,21)= 6.46, p < 0 . 0 1 , immediately postfootshock, F(2,21)=17.10, p < 0 . 0 1 , and 15 minutes later, F(2,20)=24.80, p<0.01. At each o f the 3 sampling times, blood glucose levels were greater in alloxan reactive and nonreactive animals compared to controls ( p s < 0 . 0 5 ) . Blood glucose levels increased reliably in control and diabetic rats immediately following footshock stress and remained elevated for up to 15 minutes (Fig. 3). DISCUSSION

The present findings provide support for the existence o f two distinct subgroups o f alloxan diabetic rats. One subgroup exhibited plasma E P I levels at rest which were similar to val-

FIG. 2. Plasma levels of epinephrine in control, alloxan-reactive, and alloxan-nonreactive rats under basal conditions and immediately and 15 minutes after exposure to 5 minutes of intermittent footshock. Values are presented as means and standard errors.

EZ]-CONTROL I - A L L O X A N - REACTIVE E~3-ALLOXAN- NONREACTIVE 400

-

300

-

200

-

1 O0

-

o ~

~E O

~

S [I3

niini BASAL

POST FS

+ 1 5 MIN

SAMPLING TIME

FIG. 3. Blood glucose levels in control, alloxan-reactive, and alloxannonreactive rats under basal conditions and immediately and 15 minutes after exposure to 5 minutes of intermittent footshock. Values are presented as means and standard errors.

ues for controls while the other had plasma E P I levels at rest which were elevated above controls. The reactive alloxan diabetic rats also displayed exaggerated plasma E P I responses to acute footshock stress. Interestingly, plasma NE levels under

486

LEE, KONARSKA AND McCARTY

basal conditions and following footshock stress were similar among groups. Measures of plasma catecholamines provide an accurate means of assessing the activity of the sympathetic-adrenal medullary system (11, 16, 22). Under resting conditions, plasma NE is derived almost exclusively from the sympathetic nerves. Under conditions of moderate activation, a portion of NE in blood (up to 33%) is secreted from chromaffin cells of the adrenal medulla. In animals exposed to acute stressors, as much as 45% of NE in blood is derived from the adrenal medulla. In adult rats, the exclusive source of EPI in blood is the adrenal medulla (24). The half-life of catecholamines in rat blood is approximately 70 seconds (36). In examining group differences in circulating EPI, two contributing factors should be considered- rate of secretion of EPI into blood and rate of removal of EPI from blood. In the present experiment, alloxan reactive rats maintained higher plasma concentrations of EPI but similar concentrations of NE compared to alloxan nonreactive and control rats. Similar patterns were observed in animals of the 3 groups following acute footshock stress. Compared to alloxan nonreactive and control rats, alloxan reactive rats had exaggerated plasma EPI responses but similar plasma NE responses to stress. Therefore, the higher plasma EPI levels in reactive rats appear to result from increased secretory activity of EPI-containing chromaffin cells in the adrenal medulla and not from a decreased rate of removal of EPI from blood. If the latter were involved, one would expect to observe comparable elevations in plasma NE in alloxan reactive rats as well. Alloxan reactive rats also appear to recover more slowly following acute stress as plasma EPI levels remained significantly elevated at 15 minutes postfootshock. In contrast, plasma EPI levels returned to baseline in control and nonreactive rats at 15 minutes after termination of footshock. NE-containing chromaffin cells of reactive rats may also secrete slightly more NE into blood following stress. This would explain the modest (though nonsignificant) poststress elevations in plasma NE of reactive rats (refer to Fig. 1). Reactive and nonreactive diabetic rats did not differ in the following parameters: body weight gain from treatment until the time of surgery, resting MAP and HR, or blood glucose levels before or after footshock stress. In addition, there were no dramatic behavioral differences between the two diabetic subgroups (e.g., reaction to handling, response to a novel environment, response to footshock). Elevations in plasma EPI may be attended by greater problems in diabetic control in alloxan reactive rats although this was not examined in the present experiment (7). Reactive and nonreactive diabetic rats had lower resting MAPs compared to controls. This finding is consistent with similar studies of streptozotocin diabetic rats in which MAP was measured directly in conscious animals (13,18). EXPERIMENT 2 METHOD

Animals Adult male Sprague-Dawley rats (325-350 g) were purchased from Hilltop Breeding Laboratories, Scottdale, PA. Upon arrival in our vivarium, rats were housed individually in Wahmann suspended metal cages for at least 1 week prior to experimentation. Laboratory chow and tap water were freely available at all times. The vivarium was maintained as described in Experiment 1.

TABLE 3 BODY WEIGHTS(g) OF STREPTOZOTOCIN-REACTIVE(STZ-R), STREPTOZOTOCIN-NONREACT1VE(STZ-NR)AND CONTROLRATS AT THE TIME OF STREPTOZOTOCINOR VEHICLE ADMINISTRATIONAND PRIOR TO SURGICALPLACEMENT OF TAIL ARTERYCATHETERS Group (N) Control (14) STZ-R (5) STZ-NR (3)

Pretreatment

Presurgery

399 _+ 7 391 _+ 11 403 _+ 8

397 +_ 6 360 +_ 11" 367 _+ 9*

Values are means _+ SEM for the indicated numbers of animals. *p<0.05 (paired t-test).

Streptozotocin Treatment Eight rats received intravenous (IV) injections of streptozotocin (STZ) dissolved in 5% citrate buffer (65 mg/kg in a volume of 2.5 ml/kg). Seven control rats received IV injections of 0.9% sodium chloride. Within 72 hours following STZ treatment, blood glucose levels were measured via tail clip sampling and an Accu-Chek II reflectance meter. Blood glucose levels were greater than 250 mg/dl in all STZ-treated rats. Beginning on the third day after STZ administration, rats received daily injections of regular insulin (2 U per animal) at 1700-1900 hours.

Stress Testing Animals were prepared with tail artery catheters within 5 days after administration of STZ or saline and housed as described above. Basal values of MAP and HR were measured on the day after surgery. Two days after surgery, STZ-treated animals received regular insulin (2 U per animal) at 0800 hours and basal blood samples were collected beginning at 1200 hours. Animals were then run in the acute stress protocol as described in Experiment 1 and blood samples were later assayed for plasma NE and EPI. RESULTS Our results with STZ-treated rats were quite similar to the findings presented in Experiment 1. That is, two subgroups of STZ rats were apparent based upon an analysis of plasma EPI. One group had elevated plasma levels of EPI under basal conditions and following acute footshock stress. A second group had plasma EPI levels that were similar to controls. In presenting these data, therefore, we have grouped STZ-treated rats as reactive responders and nonreactive responders. Body weights of rats at the time of administration of STZ were similar among groups, ranging from an average of 391 g for STZ reactive rats to an average of 403 g for STZ nonreactive rats. Body weights at the time of surgery (5 days after administration of STZ or saline) were reduced significantly in both of the STZ groups (ps<0.05) (Table 3). Table 4 summarizes basal values of MAP and HR for control and STZ-treated rats. MAPs were significantly different among groups, F(2,12)=4.68, p<0.05, with values for reactive and nonreactive STZ rats lower than corresponding values for controls (ps<0.05). Basal HRs were similar among groups. Basal plasma levels of NE were similar in control and diabetic rats. Immediately following footshock stress, plasma NE increased significantly for rats of the 3 groups. At 15 minutes

ACUTE STRESS IN DIABETIC RATS

487

TABLE 4

1600

BASAL VALUES OF MEAN ARTERIAL PRESSURE (MAP, mmHg) AND H E A R T RATE (HR, BEATS PER MINUTE) OF STREPTOZOTOCIN-REACTIVE (STZ-R), STREPTOZOTOCINNONREACTIVE (STZ-NR) AND C O N T R O L RATS 1 DAY AFTER SURGICAL P L A C E M E N T OF TAIL ARTERY CATHETERS

Group (N) Control (7) STZ-R (5) STZ-NR (3)

MAP 110 5:2 102 + 2 100 5:4 p<0.05

HR 324 + 12 308 5:17 316 5:16 p>0.10

E:

:I o_ i.d z_-~

ff-E

~

1400 •

F--~- CONTROL

1200

I ~-

1000

800.

z ~

600 -

u3

400 -

5

a_

STZ- REACTIVE STZ- NONREACTIVE

200-

BASAL

POST FS

+ 1 5 MIN

SAMPLING TIME

Values are means +_ SEM for the indicated numbers of animals. Levels of significance were determined by one-way ANOVAs.

postfootshock, plasma levels of NE decreased toward basal levels in control and diabetic rats (Fig. 4). Plasma levels of EPI were significantly elevated in STZ reactive rats compared to STZ nonreactive and control rats under basal conditions, F(2,12)=6.22, p<0.01. Immediately following footshock, plasma EPI levels were significantly different among groups, F(2,12)=5.94, p<0.05, with plasma EPI levels in STZ reactive rats significantly greater than values for STZ nonreactive and control rats (ps<0.05). Plasma levels of EPI at 15 minutes postfootshock returned toward basal levels and did not differ among groups (Fig. 5). Figure 6 summarizes data for blood glucose levels in STZ diabetic and control rats. Baseline blood glucose levels were significantly different among groups, F(2,12)= 11.40, p<0.01, with levels for STZ-treated rats higher than those of controls (ps<0.05). Blood glucose increased significantly in control and STZ-treated animals immediately and 15 minutes after termination of footshock (ps<0.05). Differences between control and STZ-treated rats in baseline blood glucose concentrations were maintained following footshock stress [immediately: F(2,12)=20.68, p < 0 . 0 1 , and 15 minutes postfootshock: F(2,12)=20.10, p<0.01]. DISCUSSION

As in Experiment 1, STZ diabetic rats followed a bimodal distribution with respect to levels of EPI in plasma. STZ nonreactive rats had plasma levels of EPI under basal conditions and following acute stress which were similar to controls. In contrast, STZ reactive rats had significantly higher levels of plasma EPI compared to nonreactive and control rats when at rest and immediately after footshock stress. Plasma NE at all sampling times was similar across groups. Thus, we conclude that STZ reactive rats secrete greater amounts of EPI into blood compared to rats of the other two groups. STZ reactive and nonreactive rats had lower resting MAPs compared to controls on the day after surgical placement of tail artery catheters. In addition, STZ-treated rats had similar reductions in body weight at the time of surgery and comparable levels of glucose in blood. The findings of Experiments 1 and 2 were quite similar, suggesting that alterations in EPI secretion are a common feature in approximately one-half of chemically-induced diabetic rats. Under some experimental conditions, treating STZ or alloxan diabetic rats as a homogenous group may introduce considerable variance and obscure differential treatment effects upon reactive versus nonreactive animals. This may explain some

FIG. 4. Plasma levels of norepinephrine in control, streptozotocin (STZ)-reactive, and STZ-nonreactive rats under basal conditions and immediately and 15 minutes after exposure to 5 minutes of intermittent footshock. Values are presented as means and standard errors.

1600-

U33]- CONTROL ISTZ- REACTIVE ~STZ- NONREACTIVE

14001200I

~_-n- 1000zE ~_~ 800 •

,x ~

600-

5

400-

{2200-

BASAL

P O S T FS

+ 1 5 MIN

SAMPLING TIME

FIG. 5. Plasma levelsof epinephrine in control, streptozotocin (STZ)reactive, and STZ-nonreactive rats under basal conditions and immediately and 15 minutes after exposure to 5 minutes of intermittent footshock. Values are presented as means and standard errors. inconsistencies in the literature regarding the effects of stress on blood glucose levels in diabetic rats. GENERAL DISCUSSION Stressful stimulation has been suggested to play a role in the regulation of blood glucose levels in experimental animals and in humans. The majority of animal studies to date have focused on the genetically obese mouse (C57BL/6J, ob/ob), a model of noninsulin dependent (Type II) diabetes in humans. Mice of this strain are characterized by obesity, hyperinsulinemia, insulin resistance, hyperglycemia and glucose intolerance (l). Surwit, Feinglos and their colleagues have examined the role of stress on glucoregulation in ob/ob mice. In a series of interesting experiments, they have demonstrated that ob/ob mice differ from their lean littermate controls in having exaggerated blood glucose responses to environmental stressors or exogenous administration of EPI (32-34). As mice exhibited greater increases in blood glucose following exogenous administration of EPI compared to lean littermate controls, Kuhn and coworkers concluded that ob/ob mice may have alterations in beta-adrenoceptors or in postreceptor mechanisms. They also held open the possibility that ob/ob mice may have enhanced adrenergic responses to environmental stressors (17). The BB Wistar rat has been advanced as an animal model

488

LEE, K O N A R S K A A N D M c C A R T Y 600 I~- CONTROL m l - STZ- REACTIVE F~rT-- STZ- NONREACTIVE

500 -

y~ o

400300

o

200 -

[3D 1000 BASAL

nii i

POST FS

+15

MIN

S A M P L I N G TiME

FIG. 6. Blood glucose levels in control, streptozotocin (STZ)-reactive, and STZ-nonreactive rats under basal conditions and immediately and 15 minutes after exposure to 5 minutes of intermittent footshock. Values are presented as means and standard errors.

of insulin-dependent (Type I) diabetes (26). These animals were initially recognized by causal observation of a breeding colony of Wistar rats at the BioBreeding (BB) Laboratories in Ottawa, Canada. Genetically susceptible rats are nonobese and develop acute hyperglycemia between 60 and 140 days of age (20,25). When exposed to chronic intermittent stress, the onset of diabetic symptoms is accelerated significantly in BB rats (5, 27, 28). Surwit and Feinglos (31) have advanced the hypothesis that alterations in autonomic nervous system function may play an important role in the etiology of Type II diabetes in humans. The onset of symptoms and the course of the disease in humans would be affected by an interaction between autonomic nervous system activity and environmental stress. Kemmer and co-workers (14) have examined the effects of two psychological stressors (mental arithmetic and public speaking) on cardiovascular, endocrine and metabolic responses of healthy disease-free humans and Type I diabetics in good versus poor glucoregulation. In diabetic subjects in good metabolic control (blood glucose=130+10 mg/dl), plasma EPI

responses to both stressors were greater and of longer duration compared to controls or diabetics in poor metabolic control (blood glucose=74_2 and 444± 17 mg/dl, respectively). Resting plasma EPI levels were similar in subjects of the 3 groups. These findings are quite consistent with results reported in Experiments 1 and 2 with alloxan and STZ diabetic rats, respectively. That is, Type I diabetic rats and humans exhibit exaggerated and prolonged plasma EPI responses to acute stress. In a recent study of children with I D D M , Stabler and coworkers (30) examined the effects of an acute stressor, a competitive video game, on glycemic control. They found that Type A subjects but not Type B subjects exhibited a significant glycemic response to the stressor. As a control measure, they reported that the glycemic response to consumption of a meal was similar in Type A and Type B children with IDDM. These findings are consistent with the present studies and suggest that an exaggerated plasma EPI response to stress in Type A diabetic children may precipitate the greater glycemic response to stress. In summary, approximately one-half of alloxan- and STZdiabetic rats have elevated levels of plasma EPI under resting conditions and following acute exposure to stressful stimulation. These subgroup differences in adrenal medullary secretion of EPI may have important implications for metabolic control in these animals. Alloxan- and STZ-diabetic rats may provide a valuable animal model for studies relating to the influence of sympathetic-adrenal medullary responses to stressful stimulation on metabolic control in humans with insulin dependent diabetes. ACKNOWLEDGEMENTS

This study was supported in part by a Pilot and Feasibility Grant from the University of Virginia Diabetes and Endocrine Research Center through U.S. Public Health Service Grant DK38942. Additional support was provided by ADAMHA Research Grant MH39970 and by ADAMHA Research Scientist Development Award MH00529. We thank Mr. Robert E. Stewart for technical assistance, Ms. Debbie Mundie for expert secretarial support and Drs. Daniel Cox, Robert Haynes and Joseph Lamer for support and encouragement.

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