hemodynamic effects of hypothyroidism induced by methimazole in normal and portal hypertensive rats

Digestive Diseases and Science's. VoL 40, No. 9 (September 1995), pp. 1941-1945

Hemodynamic Effects of Hypothyroidism Induced by Methimazole in Normal and

Portal Hypertensive Rats RAN OREN, MD, NIR HILZENRAT, MD, YORAM MAARAVI, MD, ARIE YAARI, MSc, and

EMANUEL SIKULER, MD

Portal hypertension is accompanied by a hyperdynamic circulatory state that shares some similarities with thyrotoxicosis. This study was conducted in order to investigate the hemodynamic effects of hypothyroidism in a rat model with portal hypertension induced by partial portal vein ligation (PVL). Four groups of 10 rats each were studied: normal control and hypothyroid rats, and PVL control and hypothyroid rats. Hypothyroidism was induced by methimazole 0.04% in drinking water. Hemodynamic measurements were performed using the radioactive microsphere technique. Induction of hypothyroidism was confirmed by elevated TSH levels. In the PVL groups, hypothyroidism ameliorated the hyperdynamic circulation. Portal venous inflow and portal pressure dropped significantly: 7.1 _+ 0.2 vs 4.8 _+ 0.3 ml/min/100 g body wt (P < 0.01) and 13.4 +_ 0.9 vs 10.9 _+ 0.8 mm Hg; (P < 0.01), respectively. In normal rats, hypothyroidism was manifested by a hypodynamic circulatory state. These results demonstrate that hypothyroidism induced by methimazole is followed by amelioration of the hyperdynamic circulation, normalization of portal venous inflow, and reduction of portal pressure.

KEY WORDS: portal hypertension: hypothyroidism: hemodynamics: methimazole; portal vein ligation.

Portal hypertension of various etiologies is accompanied by a hyperdynamic circulatory state characterized by decreased splanchnic and systemic arteriolar resistance, increased cardiac output, and increased flow into the portal system (portal venous inflow) (1-5). The clinical manifestations of the hyperdynamic circulation are warm, well-perfused extremities and bounding pulses (6, 7). Many consequences of chronic liver disease, including variceal hemorrhage

Manuscript received February 3, 1995: revised manuscript received May 2, 1995: accepted May 8, 1995.

From the Department of Medicine, Hadassah University Hospi- tal, Mount Scopus, Jerusalem: and Hepatic Hemodynamic Llbo- ratory, Department of Medicine, Soroka Medical Center and Fac- ulty of Health Sciences, Ben-Gurion University of the Negev, Beer-Sheva, Israel.

Address for reprint requests: Dr. Emanuel Sikuler, Department of Medicine B', Soroka Medical Center, POB 151, Beer-Sheva, 84101 Israel.

and ascites, may be tied intimately to the presence of these hemodynamic alterations and nearly all clini- cally useful drugs that reduce portal pressure act by ameliorating the hyperdynamic circulatory state (7-9).

Hyperthyroidism is another hyperdynamic circulatory state also manifested by warm, well-perfused extremities and bounding pulses; therefore, it shares some similarities with portal hypertension. Proprano- lol, a drug used to reduce portal pressure is used for the treatment of cardiovascular expressions of thyrotoxicosis as well (8-10). Induction of hypothyroidism may therefore serve as a potential means to amelio- rate the hyperdynamic circulation and thus to reduce portal pressure in portal hypertensive subjects. More- over, propylthiouracil (PTU), an antithyroid drug, was suggested as a possible therapy for alcoholic liver

Digestive Diseases and Sciences. Vol. 4(1. No. 9 (September 1995] Ill 63- 21 I h/95,'1190(I- I t?41 $117.511jll ~ 1995 P[e n u m Pub l i sh ing C o r p o r a t i o n

1941

OREN ET AL

disease and was also found to have a pro tec t ive effect against ga l ac to samine - induced hepa toce l lu l a r necro- sis (11-13) .

This s tudy was conduc t ed in o r d e r to invest igate the h e m o d y n a m i c effects of pha rmaco log ica l ly induced hypothyro id i sm in a por ta l hyper tens ive rat model .

M A T E R I A L S AND METHODS

Forty male Sprague-Dawley rats (Charles River Co., Kent, UK) weighing 220-240 g were used. Portal hypertension was induced in 20 rats. The remainder served as normal controls. The operative procedure for production of portal hypertension has been previously described (2). In brief, the animals were anesthetized with ether and ketamine HCI (150 mg/kg intramuscularly). The portal vein was isolated and stenosis was achieved by a single ligature of 3-0 silk around the portal vein and a 20-gauge blunt- tipped needle. The needle was then removed, the portal vein was allowed to reexpand distal to the stenosis, and the abdomen was closed by layers. After the operation, the animals were housed in plastic cages and allowed free access to rat chow and water until the time of the hemodynamic studies. Normal, not sham-operated, rats were used in this study as controis, since in our experience the hemodynamics over a wide range of physiological stimuli and the response to methimazole in normal and sham-operated rats are identical (unpublished data). The animals were kept under veterinary care in the research animal facility of the Faculty of Healthy Sciences, Ben Gurion University, and treated according to APS Guiding Principles for Research Involving Animals.

Hypothyroidism was induced by administrat ion of methimazole (Taro-Israel) in drinking water at a concen- tration of 0.04% for 14 days, after one week of access to tap water (14). The rats were divided into four groups of 10 each: group 1--normal rats drank tap water for three weeks [normal water (NW)]; group 2--normal rats with hypothyroidism induced after one week of drinking tap water [normal hypothyroid (NH)]; group 3--portal hypertensive control rats were studied three weeks after partial portal vein ligation (PVL), while drinking tap water [portal vein ligation-water (PVL-W)]; and group 4- -por ta l hypertensive rats with hypothyroidism induced beginning at day 8 after PVL [portal vein ligated-hypothyroid (PVL-H)].

Hemodynamic studies were performed using the radioactive microsphere technique. The technique used for the hemodynamic measurements has been detailed previously (2-4). Animals were anesthetized with ketamine HC1 (150 mg/kg intramuscularly). The left femoral and right carotid arteries were exposed and cannulated with PE-50 catheters, which were connected to a Statham p-23-Db strain-gauge transducer. Permanent recording of arterial pressure was made on a Gilson biscriptual polygraph, model ICM-8, inscription recorder (Gilson Medical Electronics Inc., Middleton, Wisconsin). The right carotid artery catheter was advanced into the left ventricle during continuous pressure monitoring. Body temperature was maintained at 37.0 -+ 0.5~ (rectal) by a heating lamp.

Cardiac output and regional organ blood flow were measured using a reference sample method (15). The reference sample was withdrawn from the left femoral artery catheter into a preweighed syringe for 75 sec at an approximate rate of 1.0 ml/min using a Harvard pump (Harvard Apparatus, Millis, Massachusetts). Fifteen seconds after beginning withdrawal of the reference sample, approximately 60,000 polymeric resin, t~ microspheres (16.5 _+ 0.1 /xm diameter) were injected into the left ventricle over a period of 10-15 sec. The catheter was flushed with 0.2 ml of 0.9% NaC1. A volume of 0.9% NaC1, equal to the reference sample, was reinfused, and the arterial blood pressure mon- itored to ensure a stable preparation. Splenic pulp pressure, an indirect measurement of portal pressure, was used in this study to measure portal pressure. A small parasagittal inci- sion (1.5-2.0 cm) was made in the left upper abdomen through the skin and muscle. The spleen was exposed by retraction of the perisplenic fat with care taken to avoid undue manipulation. Intrasplenic pressure was measured by inserting into the splenic parenchyma a fluid-filled 19-gauge needle connected by 6-cm-long tubing to a Statham strain- gauge transducer calibrated for venous pressure. The exter- nal zero reference point was set at the midportion of the animal. The pressure reading was considered acceptable when a stable recording was produced and normal respira- tory variations were observed. Hemostasis was obtained after needle removal by applying cyanoacrylate glue (Poly- glue-Potytoff, Israel) to the site of insertion.

Blood samples for hormone assay were withdrawn from the femoral catheter at the end of the experiment. The blood was centrifugated and the serum kept at -20~ until assayed.

The animals were killed at the end of the experiment by a bolus injection of KC1. The abdominal organs, kidneys, lungs, and testes were dissected and weighed. The radioactivity (cpm) of each organ was determined in an autogamma scintillation counter (Packard, Downers Grove, Illinois). Kidney and testicular radioactivity was determined to check for adequate mixing of microspheres. The larger organs were dissected into smaller portions for uniform geometry within the scintillation counting tubes.

Cardiac output (CO) and regional organ blood flow (OBF) were calculated after the ")3Ru injection as follows (4):

CO(ml/min)

injected radioactivity (cpm) • reference blood flow rate (ml/min)

= reference blood radioactivity (cpm)

OBF(ml/min)

organ radioactivity (cpm) x reference blood flow rate (ml/min)

- reference blood radioactivity (cpm)

At least 300 microspheres had to be trapped in both reference sample and organs to ensure the validity of the measurements (15).

Cardiac index was calculated as cardiac output per 100 g body weight.

The portal venous inflow (PVI) was the sum of the blood

1942 Digestive Diseases and Sciences, Vol. 40, No. 9 (September 1995)

HYPOTHYROIDISM IN PVL RATS

3.0

2.5

2.0

E 1.5

D

1.0

0.5

0.0

I [ N - W N - H

PVL - W

1 PVL - H

I I

Fig l. TSH levels in untreated and methimazole-treated normal and portal hypertensive rats. N-W, normal water; N-H, normal hypothyroid; PVL-W, portal vein ligated, water; PVL-H, portal vein ligated, hypothyroid. *P < 0.01 versus respective untreated group.

flow to stomach, spleen, small and large intestines, pan- creas, and mesentery. This calculation represents total blood flow into the portal venous system. For practical reasons, flows were all expressed in milliliters per minute per 100 g body weight. This was done in order to avoid variations in flow measurements that might be due to small discrepancies in weight between animal groups.

Vascular resistance (R) in various vascular systems was calculated in peripheral resistance units according to the following equation:

&P (mm Hg) R (mm Hg*/min/ml) Q (ml/min)

In calculating total peripheral resistance and renal resistance, AP equals mean atrial pressure minus right arterial pressure and Q is the cardiac output and renal blood flow, respectively. The right atrial pressure in rats, measured in previous studies, was found to be less than 1 mm Hg (16) and therefore was disregarded. In calculating splanchnic arteriolar resistance, AP equals mean arterial pressure minus portal pressure, and Q is the portal venous inflow.

Serum concentrations of TSH and T 3 were determined using a chemiluminescent immunoassay. (Immulite, DPC). We measured T 3 since it is the active hormone.

Results are expressed as means - SE. Statistical analysis was performed by one way analysis of variance, and the Newman-Keuls correction of multiple comparisons. Results were considered significant at P < 0.05.

RESULTS

At the time of the study, the body weight of the four groups was similar (NW, 368 _+ 20; NH, 348 +_ 24; PVL-W 350 _ 31; PVL-H 338 +_ 30 g).

T 3 and TSH Levels (Figure 1). T 3 levels in all groups were not significantly different (NW, 1.33 _+ 0.10; NH, 1.17 _ 0.10; PVL-W, 1.30 _+ 0.05; PVL-H, 1.10 _ 0.10 mmol/liter). The TSH levels of the two methimazole-treated groups (1.84 +_ 0.60 and 1.47 _ 0.50 mmol/liter, NH and PVL-H, respectively; P = NS) were significantly higher than those of the untreated ones (0.20 _+ 0.06 and 0.13 _+ 0.08 mmol/liter, NW and PVL-W, respectively; P = NS) (P < 0.01 vs the respective untreated group).

Untreated portal hypertensive rats (PVL-W) had a systemic hyperdynamic circulation, as manifested by increased cardiac index and renal blood flow, and decreased peripheral resistance. Hepat ic arterial blood flow in these rats was also elevated (Table 1). Treatment with methimazole resulted in a reduction in heart rate and cardiac index in both normal and PVL rats. Mean arterial pressure and muscular blood flow also decreased as a result of methimazole treatment, although only in the normal rats was the change statistically significant. A reduced renal blood flow following methimazole administration was observed only in the portal hypertensive rats. Hepatic arterial blood flow decreased significantly only in normal rats.

TABLE 1. HEMODYNAMIC DATA*

N W P N H PVL- W P P V L - H

HR (bpm) 339 _+ 12 <0.001 267 _+ 9 351 _ 13 <0.001 270 -+ 11 MAP (ram Hg) 105.4 + 4.0 <0.05 85.2 _+ 3.8 97.8 _ 5.5 NS 85.8 -+ 7.2 CI (ml/min/100 g body wt) 38.8 -- 1.6 <0.01 28.2 --- 2.0 49.0 _ 3.5t <0.01 37.8 _+ 2.4.]: TPR (mm Hg*min*100 g body

wt/ml) 2.74 __ 0.09 NS 3.12 ___ 0.21 2.02 _+ 0.19t NS 2.37 - 0.25:1: RBF (ml/min/100 g body wt) 3.50 _+ 0.20 NS 3.32 _+ 0.19 4.54 +_ 0.37t <0.01 3.17 -+ 0.24 RR (mm Hg/min/ml/100 g

body wt) 31.0 - 2.0 NS 26.0 --- 0.8 23.8 _+ 3.3 NS 29.0 +- 4.2 MBF (ml/min/g) 0.08 -+ 0.01 <0.05 0.04 _+ 0.01 0.10 - 0.01 NS 0.07 -+ 0.01 HABF (ml/min/100 g body wt) 0.63 -+ 0.10 <0.05 0.38 _+ 0.07 1.03 - 0.21t NS 0.68 -+ 0.12~

*HR, heart rate; MAP, mean arterial pressure; CI, cardiac index; TPR, total peripheral resistance; RBF, renal blood flow; RR, renal resistance; MBF, muscular blood flow; HABF, hepatic arterial blood flow; NW, normal water; NH, normal hypothyroid; PVL-W, portal vein ligated, water; and PVL-H, portal vein ligated, hypothyroid.

t P < 0.05 vs NW. P < 0.05 vs NH.

Digestive Diseases and Sciences, Vol. 40, No. 9 (September 1995) 1943

OREN ET AL

2 0

18

16

14

0"1 12 '-I-

E lo E 8

6

4

2

0

F i g 2. Por ta l P V L W : " * P

N - W

[ 7 ~ N - H

PVL - W

I I PVL - H

pres su re . A b b r e v i a t i o n s as in F igure 1. *P < 0.01 vs < 0.06 vs N H .

Portal pressure (Figure 2) in the PVL-W group was significantly higher than in the normal control: 13.4 +_ 0.9 vs 8.1 _+ 0.4 mm Hg (P < 0.01). In the portal hypertensive rats, a significant reduction of portal pressure to 10.9 +_ 0.8 mm Hg was observed in the methimazole-treated group (P < 0.01). In the normal rats, portal pressure decreased to 6.2 _+ 0.4 mm Hg, after methimazole administration. However, this decrease was not significant (P < 0.06).

Portal venous inflow (Figure 3) in the PVL-W rats was significantly higher than in the N-W rats (7.1 _+ 0.2 vs 5.2 + 0.4 ml/min/100 g body wt; P < 0.01). Portal venous inflow was significantly reduced in both the PVL-H group (4.8 m 0.3 ml/min/100 g body wt: P < 0.01) and in the N-H group (3.7 -+ 0.4 ml/min/100 g body wt; P < 0.01) as compared to that of the PVL-W and N-W groups, respectively. The portal venous inflow in the PVL-H group was not significantly different from that of the N-W group.

Portal venous inflow, expressed as percentage of cardiac output, was similar in all four groups.

12

10

[ i H - W r-J - H

I PVL - H c6

0 0

E 4

0 _ _ I

Fig 3. Por ta l v e n o u s inflow (PVI) . A b b r e v i a t i o n s as in F igure 1. *P < 0.01 vs the respec t ive e u t h y r o i d g roups ; **P < 0.01 vs N W group.

Changes in blood flow to the individual organs that drain into the portal system were parallel to portal venous inflow changes in all groups, although these changes did not always reach statistical significance.

Splanchnic arteriolar resistance was not significantly altered by methimazole: 19,9 _ 2.3 vs 23,4 _+ 2.1 mm Hg*min*100 g body wt/ml in NW and NH, respectively, and 12.1 _+ 0.9 vs 15.9 -+ 0.6 mm Hg*min*100 g body wt/ml in PVL-W and PVL-H, respectively. However, in both PVL-W and PVL-H, the values obtained were significantly lower (P < 0.05) than in the N-W and N-H groups.

DISCUSSION

Administration of methimazole to normal and portal hypertensive rats led to hypothyroidism, as indi- cated by elevated TSH and reduced heart rate. This was followed by marked hemodynamic changes: In portal hypertensive rats, hypothyroidism ameliorated the hyperdynamic circulatory state. Cardiac output and portal venous inflow were significantly reduced, returning to normal levels. Portal pressure was also significantly decreased. In normal rats a hypodynamic circulatory state was observed. This was characterized by reduced heart rate, cardiac output, and splanchnic blood flow.

Methimazole and not propylthiouracil (PTU) was chosen in order to induce hypothyroidism in our study since acute administration of PTU was found to increase portal blood flow through an effect indepen- dent of its thyroid action (17). Acute administration of methimazole, a drug that has the antithyroid action of PTU, does not affect portal blood flow (17). There- fore, the hemodynamic alterations observed in this study are due to hypothyroidism and not a direct effect of the drug.

Renal blood flow (RBF) in the normal rats was not modified by methimazole treatment. On the other hand, the elevated RBF observed in the untreated PVL rats was decreased by methimazole treatment. These results suggest that the hemodynamic effects of hypothyroidism may be influenced by the pretreat- ment basic hemodynamic state.

The hyperdynamic circulation of portal hypertension is well established. Nevertheless, the exact mech- anism responsible for these hemodynamic changes is still undefined (1). Although /3-adrenergic blockers are not specific inhibitors of the hyperdynamic circulation of portal hypertension, they are a logical therapeutic approach for this hemodynamic abnormality (8). Hyperthyroidism is also a hyperdynamic circu-

] 944 Digestive Diseases and Sciences. l'~)l. 40. Nu. 9 r 19951

HYPOTHYROIDISM IN PVL RATS

latory state (18) and shares many similarities with portal hypertension, eg, high cardiac output, low peripheral resistance, and well-perfused extremities. Hyperthyroidism is associated with excess catechol- amine activity (19). Moreover thyroid hormones po- tentiate the action of catecholamines (20). Proprano- 1ol is useful for treatment of the cardiovascular expressions of thyrotoxicosis (18) and can also inhibit the peripheral conversion of T4 to T 3 (10). Our results, which demonstrate amelioration of the hyperdynamic circulation of portal hypertension by induction of hypothyroidism, provide another similarity between portal hypertension and hyperthyroidism. The hemodynamic changes in our portal hypertensive rats were similar to those observed in portal vein- ligated rats treated with /3-blockers in a dose equiv- alent to doses used in patients with liver cirrhosis (21) (in whom a 25% decrease in heart rate is gen- erally used as an approximate index of therapeutic /3-blockade).

In summary, our results demonstrate that hypothyroidism, induced by methimazole, ameliorates the hyperdynamic circulatory state and reduces portal pressure in portal hypertensive rats. Although caution should be used in extrapolating results from animal models to humans, this form of treatment may have potential for treating portal hypertensive chronic liver disease. Moreover, antithyroid drugs have also been shown to have beneficial effects on parenchymal liver damage (22); therefore, we suggest that the exact role of this therapeutic modality in humans will be evalu- ated in clinical trials.

REFERENCES

1. Mahl TC, Groszmann R J: Pathophysiology of portal hypertension and varicea[ bleeding. Surg Clin North Am 70:251-266, 1990

2. Vorobioff J, Bredfeldt JE, Groszmann R J: Hyperdynamic circulation in a portal hypertensive rat model: A primary factor for maintenance of chronic portal hypertension. Am J Physiol 244(Gastrointest Liver Physiol 7):G52-G57, 1983

3. Vorobioff J, Bredfeldt JE, Groszmann RJ: Increased blood flow through the portal system in cirrhotic rats. Gastroenter- ology 87:1120-1126, 1984

4. Sikuler E, Buchs AE, Yaari A, Keynan A: Hemodynamic characterization of conscious and ketamine-anesthetized bile duct ligated rats. Am J Physiol 260(Gastrointest Liver Physiol 23):G161-G166, 1991

5. Bosch J, Enriquez R, Groszmann R J, Storer EH: Chronic bile duct ligation in the dog: Hemodynamic characterization of a portal hypertensive model. Hepatology 3:1002-1007, 1983

6. Murray JF, Dawson AM, Sherlock S: Circulatory changes in chronic liver disease. Am J Med 24:358-367, 1958

7. Kowalski H J, Adelmann AB: The cardiac output at rest and in Laennec's cirrhosis. J Clin Invest 32:1025-1033, 1953

8. Sikuler E:/3-Adrenergic blockers for portal hypertension: Les- sons from animal models. J Hepatol 12:133-135, 1991

9. Kroeger RJ, Groszmann R J: Effect of selective blockade of/32 adrenergic receptors on portal and systemic hemodynamics in a portal hypertensive rat model. Gastroenterology 88:896-900, 1985

10. Wortofsky L, Ingbar SH: In Principles of Internal Medicine, Vol 2, 12th ed. New York, McGraw-Hill, 1991, p 1706

11. Orrego H, Blake JE, Blendis LM, Comptom KV, Israel Y: Long-term treatment of alcoholic liver disease with propylthiouracil. N Engl J Med 317:1421-1427, 1987

12. Carmichael FJ, Orrego H, Saldivia V, Israel Y: Effect of propylthiouracil on the ethanol-induced increase in liver oxy- gen consumption in awake rats. Hepatology 18:415-421, 1993

13. Schwedes U, Wdowinski JM, Siede WH, Vardarli I, Schmidt R, Usadel K/I: Effect of hypometabolism on cell injury. Klin Wochenschr 64(suppl VII):146-148, 1986

14. Abelenda M, Puerta ML: Cold-induced thermogenesis in hypothyroid rats. Pfleugers Arch 416:663-666, 1990

15. Ishise S, Pegram BL, Yamamoto J, Kitamura Y, Frohlich ED: Reference sample microsphere method: Cardiac output and blood flows in conscious rats. Am J Physiol 239(Heart Circ Physiol 8):H443-H449, 1980

16. Sikuler E, Groszmann R J: Interaction of flow and resistance in maintenance of portal hypertension in a rat model. Am J Physiol 250(Gastrointest Liver Physiol 13):G205-G212, 1986

17. Kawasaki T, Carmichael FJ, Giles G, Saldivia V, Israel Y. Orrego H: Effects of propylthiouracil and methimazole on splanchnic hemodynamics in awake and unrestrained rats. Hepatology 10:273-278, 1989

18. Skelton CL: The heart and hyperthyroidism. N Engl J Med 307:1206-1207, 1982

19. Morkin E, Flink IL, Goldman S: Biochemical and physiologic effects of thyroid hormone on cardiac performance. Prog Car- diovasc Dis 25:435-465, 1983

20. Williams LT, Lefkowitz RJ, Watanbe AM: Thyroid hormon regulation of beta adrenergic receptor number. J Biol Chem 252:2787-2789, 1977

21. Lee SS, Hadengue A, Girod C, Braillon A, Lebrec D: Diver- gent circulatory effects of betaxolol in eoncious and anesthetized normal and portal hypertensive rats. J Hepatol 12:157- 161, 1991

22. Orrego H, Blake JE, Blendis LM, Compton KV, Volpe R, Israel Y: Long-term treatment of alcoholic liver disease with propylthiouracil. Part 2: Influence of drop-out rates and of continued alcohol consumption in a clinical trial. J Hepatol 20:343-349, 1994

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hemodynamic effects of hypothyroidism induced by methimazole in normal and portal hypertensive rats

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