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Effect of Caffeine on Thyroid and Pituitary Function in Newborn Rats (40)

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Effect of Caffeine on Thyroid and Pituitary Function in Newborn Rats (40) Pediatr. Res. 17: 592-595 (1983) Effect of Caffeine on Thyroid and Pituitary Function in Newborn Rats (40) MARTINE CLOZEL, CHARLOTTE L. BRANCHAUD, GLORIA S. TANNENBAUM, JEAN H. DUSSAULT, AND JACOB V. ARANDAC4" Developmental Pharmacology and Perinatal Research Unit, Department of Endocrinology, McGiN University- Montreal Children's Hospital Research Institute, Montreal, Quebec, Canada and Department of Endocrinology, Universite Laval, Quebec City, Quebec, Canada Summary experiments were respectively designed to evaluate the acute and the chronic effects of caffeine. Each set used three groups: a The possibility that caffeine, a central nervous system stimulant control group, which received saline; a low dose caffeine group, used in neonatal apnea, may produce acute or chronic changes in which received 5 mg/kg of caffeine dissolved in saline; and a high growth hormone (GH), thyroxine (T4) and thyrotropin (TSH) was dose caffeine group, which received 50 mg/kg. Caffeine (37) or studied in the newborn rat. Five-day-old rats were separated into saline was injected intraperitoneally in a volume of 0.05 ml/rat. three groups: control (0) group receiving saline, Group I (low dose Each group consisted of 30-35 rats. In the acute experiment, the caffeine) receiving 5 mg/kg and Group I1 (high dose caffeine) animals were sacrificed at various time intervals after a single receiving 50 mg/kg. Acute effects were studied at 2, 4, and 24 h injection of saline or caffeine: 2, 4, and 24 h. In the chronic after injection. Chronic effects were studied 24 h after the last of experiment, animals were injected daily for a 10 day period. They 10 daily injections. GH, T4, and TSH were measured by radioim- were sacrificed 24 h after the last injection. The rat pups were munoassay and caffeine by high pressure liquid chromatograph. kept with their suckling mothers throughout the experiments and GH was increased at all times and all doses after a single injection maintained on a 12 h light-dark cycle. of caffeine. After chronic therapy, the increase in GH was small, suggesting depletion of pituitary reserve. A high dose of caffeine At the end of the 24 h experiment, several rats (n = 6) of the had a biphasic effect on T4 with an increase at 4 h and a decrease control group and of the low dose caffeine group received TRH, at 24 h. Thyrotropin-releasing hormone (TRH)-induced TSH in order to evaluate the effect of caffeine on TRH-induced TSH release at 24 h was not influenced by caffeine administration. release. In the chronic experiment, each of the three groups was Chronic caffeine therapy stimulated both T4 and TSH; however, divided into two: for the measurement of either basal TSH (n = TRH-stimulated TSH release was decreased, suggesting that 20) or TRH-stimulated TSH (n = 10). Rats receiving TRH (38) chronic therapy may blunt pituitary TSH response. were injected intraperitoneally with 100 ng TRH/100 g body weight, 15 min before sacrifice, as described by Walker et al. (34). Abbreviations TRH was dissolved in saline. The volume of TRH in saline was 0.05 m1/10 g of body weight given intraperitoneally. GH, growth hormone At the termination of all studies, animals were sacrificed by PDE, phosphodiesterase rapid decapitation at the same time of the day, between 14:OO and T3, triiodothyronine 14:30. Blood was collected, and plasma or serum obtained was T4, thyroxine stored at -20°C until subsequent analysis. T4 and GH were TRH, thyrotropin releasing hormone measured in plasma, TSH in serum. T4 was measured using an TSH, thyrotropin Amerlex T4 radioimrnunoassay kit (Amersham Corporation), TSH (34) and GH (33) by double antibody radioimmunoassay with Besides the stimulant effect of the methylxanthines on the materials supplied by the NIADDK (39). The TSH intraassay central nervous system and on cardiac inotropy and chronotropy, variation is 6-8%; interassay variation is 12% and the sensitivity these compounds exert a variety of hormonal and metabolic was between 10-15 ng per tube. For the GH assay, the intra- and effects. The administration of two methylxanthines, caffeine and interassay coefficients of variation determined for a sample mean theophylline, to adult rats results in decreased GH, TSH, TQand of 25 ng/ml were 8.9 and 7.2, respectively. The minimum detect- thyroxine T4 serum levels (29). But in in vitro isolated organ able concentration in the radioimmunoassay defined as the con- preparations, caffeine increases T4 (3) and TSH (28) release, and centration of rat GH, which resulted in a binding of ['251] or GH theophylline increases the release of GH (32). Published data on to antiserum that was 2 standard deviations below the mean the hormonal effects of methylxanthines in humans are contradic- binding achieved in the absence of GH, was 3.1 ng/ml. tory. Peracchi et al. (25) and Ensinck et al. (17) observed a decrease Caffeine and one of its potential metabolites, theophylline, were in GH level in acromegalic patients after aminophylline. In normal measured by high pressure liquid chromatography (2). The sen- men, Blackard et al. (8) and Peracchi et al. (25) did not find any sitivity of the caffeine assay was 0.1 mg/liter and the coefficient modification in basal GH after aminophylline, whereas, Ensinck of variance at 10 mg/liter was 4.0%. et al. (17) described a decrease in GH serum level. After a low Statistical analysis was performed using two-way analysis of dose of oral caffeine given to adult healthy men, neither GH, TSH variance (36) or unpaired Student's t test. All values are reported nor Tg were modified (30). Because caffeine is currently used in as mean k S.E. the premature neonate for the treatment of apnea, we evaluated the changes in GH, T4, and TSH after caffeine in the newborn RESULTS rat. GH. Figure 1 shows the effects of caffeine on plasma GH levels MATERIALS AND METHODS in the short term experiment. There was a trend to an increase in plasma GH levels in response to both doses of caffeine at all time Five-day-old male Sprague-Dawley rats were obtained from the points examined. The effect increased significantly with the dose. Canadian Breeding Farm, St. Constant, Quebec. Two sets of After chronic treatment (Fig. 2) a significant increase (P < 0.05) THYROID AND PITUITARY FUNCTION 593 .. ured 24 h after either saline (control group) or 50 mg/kg of caffeine (treated group) in the 4-day-old rats. No significant change in TRH-stimulated TSH was noted (377.8 f 32.5 versus 360.7 f 37 pg/100 ml). Table 1 shows the effect of chronic caffeine treatment on basal and TRH-stimulated T4 and on basal and TRH-stimulated TSH. Both low and high doses of caffeine significantly (P < 0.001) - increased Tq levels (5.06 & 0.29 and 6.04 f 0.22, respectively, E .-_I versus 4.12 f 0.27 &dl in 0 dose). The same pattern was found E? tw - after TRH administration but there was no significant difference a= a in each group between basal T4and TRH-stimulated T,, suggest- 5a ing that the 15 min interval between TRH administration and n sacrifice was probably too short to produce changes in T4 concen- h tration. Chronic therapy with caffeine resulted in T4 levels greater then control animals (Table l), suggesting an escape phenomenon from the biphasic effect obtained with acute caffeine administra- tion. Basal TSH was slightly increased by a low dose (17.9 rt: 2.57 f r: versus 10.6 1.43 &dl) and significantly by a high dose of f Tlrw Arler Inlcc:lon (Hrs] caffeine (25.1 6.93 versus 10.6 + 1.43 &dl, P < 0.05) (Table 1). The TSH levels after TRH stimulation was considerably greater Fig. I. Effect of a single administration of saline (n = lo), caffeine 5 than basal levels (216.6 f 45.5 versus 10.6 + 1.4 pg/lOO ml); (n = lo), and caffeine 50 mg/kg (n = 9) on growth hormone (GH) however, chronic caffeine therapy with both low (5 mg/kg) and plasma levels (mean _t S.E.) at different times after injection. *P< 0.05 high doses (50 mg/kg) attenuated the TSH response to TRH and ***P< 0.001, compared to the control group (saline). stimulation. In the caffeine-treated rats, the TSH levels in the TRH-stimulated rats were much higher than the basal TSH. But TSH levels after TRH stimulation in the low and high caffeine- treated rats were significantly lower compared to the non-caffeine treated group (Table I), suggesting that chronic caffeine therapy may possibly lead to exhaustion of pituitary reserve. 1j@ Caffeine assay. The plasma levels of caffeine and of theophylline obtained after injection of caffeine are shown in Table 2. The 110 peak plasma concentration of caffeine after 5 mg/kg is 5.26 f 0.38 - mg/liter, which is within the lower therapeutic range for the -c -_I management of neonatal apnea. After 50 mg/kg, the peak con- CD L centration is 67.57 * 2.84 mg/liter, which is supra-therapeutic 5 lw concentration. The caffeine level 24 h after the last injection of a chronic u,2 a treatment with 50 mg/kg (2.00 f 1.26 mg/liter) is much lower L than 24 h after a single injection of 50 mg/kg (20.65 f 3.36 mg/ liter), suggesting that there is no accumulation of the drug and/or that the clearance is greater at 15 days of age than at 5 days. No demethylation of caffeine into theophylline was detected after a n single injection of 5 mg/kg.
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