Adenosine A3 receptors regulate heart rate,motor activity and body temperature

Aim: To examine the phenotype of mice that lack the adenosine A₃ receptor (A₃R). Methods: We examined the heart rate, body temperature and locomotion continuously by telemetry over several days. In addition, the effect of the adenosine analogue R-N⁶-phenylisopropyl-adenosine (R-PIA) was examined. We also examined heat production and food intake. Results: We found that the marked diurnal variation in activity, heart rate and body temperature, with markedly higher values at night than during day time, was reduced in the A₃R knock-out mice. Surprisingly, the reduction in heart rate, activity and body temperature seen after injection of R-PIA in wild type mice was virtually eliminated in the A₃R knock-out mice. The marked reduction in activity was associated with a decreased heat production, as expected. However, the A₃R knock-out mice, surprisingly, had a higher food intake but no difference in body weight compared to wild type mice. Conclusions: The mice lacking adenosine A₃ receptors exhibit a surprisingly clear phenotype with changes in diurnal rhythm and temperature regulation. Whether these effects are due to a physiological role of A₃ receptors in these processes or whether they represent a role in development remains to be elucidated.     

Eliminating the antilipolytic adenosine A1 receptor does not lead to compensatory changes in the antilipolytic actions of PGE2 and nicotinic acid

Aim: We examined whether compensatory changes after adenosine A₁ receptor knockout [A₁R (−/−)] eliminate the antilipolytic actions mediated by this receptor. Methods: Lipolysis experiments were performed on adipocytes prepared from the wild type A₁R (+/+), A₁R (−/−) and heterozygous mice. Gene expression was assayed with cDNA microarray technique and real‐time PCR; protein expression with immunoblotting. Results: The A₁R was the only adenosine receptor involved in lipolysis. The effects of adenosine deaminase and 2‐chloroadenosine were abolished in A₁R (−/−) mice. The IC₅₀ value of 2‐chloroadenosine doubled from 16.6 to 33.6 nm when half of the A₁Rs were eliminated. Adrenergic α₂ agonists had no effects on lipolysis. Prostaglandin E₂ (PGE₂) inhibited lipolysis with an IC₅₀ value of 5.8 nm (4.7–7.2 nm ) in the A₁R (+/+) mice and 10.6 nm (9.0–12.6 nm ) in the A₁R (−/−) mice. Nicotinic acid inhibited lipolysis with an IC₅₀ value of 0.30 μm (0.19–0.46 μm ) in the A₁R (+/+) mice and 0.24 μm (0.16–0.37 μm ) in the A₁R (−/−) mice. G_iα₁ mRNA was significantly up‐regulated in adipose tissue from A₁R (−/−) mice. However, immunoblotting showed that G_iα1 was not up‐regulated at the protein level. Conclusion: The A₁R mediates the antilipolytic actions of adenosine. Deletion of the A₁R in mice does not result in compensatory increases in G‐protein‐mediated antilipolytic actions of PGE₂ or nicotinic acid.     

Decreased behavioral activation following caffeine, amphetamine and darkness in A3 adenosine receptor knock-out mice

We have examined behavioral consequences of genetic deletion of the adenosine A₃ receptors in mice. The open field behavior of A₃ adenosine receptor knock-out (A₃R KO) mice was investigated both under basal conditions and after stimulation with psychostimulants. Adolescent (21 day-old) and adult A₃R KO males showed an increase in overall motor activity compared to wild type (WT) males, but the type of activity differed. The motor activity, especially rearing, was also higher in A₃R KO compared to WT adult females. A₃ receptors have a low affinity for caffeine and it was therefore surprising to find a decreased response to stimulation with either caffeine or amphetamine in A₃R KO as compared to WT mice in males as well as females. Telemetry recordings also showed a significantly smaller increase in activity upon darkness in A₃R KO. There were no compensatory changes in the mRNA expression of any other adenosine receptor subtypes (A₁, A_2A and A_2B) or any changes in dopamine D₁ and D₂ receptor binding in A₃R KO brains. Challenge with the developmental toxicant methylmercury (1 µM in drinking water) during pregnancy and lactation did not cause any behavioral alterations in adolescent and adult WT female offspring. In contrast, the A₃R KO female offspring displayed changes in locomotion indicating an interaction between perinatal methylmercury and adenosine A₃ receptors. In conclusion, despite low expression of A₃ receptors in wild type mouse brain we observed several behavioral consequences of genetic elimination of the adenosine A₃ receptors. The possibility that this is due to a role of A₃ receptors in development is discussed.     

Oral administration of Japanese sake yeast (Saccharomyces cerevisiae sake) promotes non‐rapid eye movement sleep in mice via adenosine A2A receptors

We have demonstrated previously that Japanese sake yeast improves sleep quality in humans. In the present study, we examined the molecular mechanisms of sake yeast to induce sleep by monitoring locomotor activity, electromyogram and electroencephalogram in mice. Oral administration of Japanese sake yeast (100, 200, and 300 mg kg^?1) decreased the locomotor activity by 18, 46 and 59% and increased the amount of non‐rapid eye movement (NREM) sleep by 1.5‐, 2.3‐ and 2.4‐fold (to 37 ± 6, 57 ± 8, and 60 ± 4 min from 25 ± 6 min in the vehicle‐administered group, respectively) in a dose‐dependent manner for 4 h after oral administration. However, Japanese sake yeast did not change the amount of rapid eye movement (REM) sleep, the electroencephalogram power density during NREM sleep or show any adverse effects, such as rebound of insomnia, during 24 h postadministration and on the next day. An intraperitoneal pretreatment with an adenosine A_2A receptor‐selective antagonist, ZM241385 (15 mg kg^?1), reduced the amount of NREM sleep of sake yeast‐administered mice to the basal level, without changing basal amount of sleep. Conversely, an A₁ receptor‐selective antagonist, 8‐cyclopentyltheophylline (10 mg kg^?1), did not affect the sleep‐promoting effect of Japanese sake yeast. Thus, Japanese sake yeast promotes NREM sleep via activation of adenosine A_2A but not A₁ receptors.     

Caffeine reverses antinociception by amitriptyline in wild type mice but not in those lacking adenosine A1 receptors

Amitriptyline is used to treat neuropathic pain in humans. It produces antinociception in several animal models of pain, and this effect is blocked by methylxanthine adenosine receptor antagonists which implicates adenosine it its actions. Here, the antinociceptive effect of amitriptyline, and the ability of caffeine to reverse it, were examined using the formalin test (a model of persistent pain) in wild type mice and mice lacking the adenosine A₁ receptor (A1R). Amitriptyline produced dose-related suppression of flinching in wild type mice following both systemic and intraplantar drug administration; both of these effects were unaltered in A1R −/− mice. Following systemic administration, caffeine reversed the systemic effect of amitriptyline in wild type, but not A1R −/− mice; −/+ mice exhibited an intermediate effect. Intraplantar administration of caffeine also reversed the effect of intraplantar amitriptyline in A1R +/+, but not in −/− or +/− mice. These results indicate that adenosine A₁ receptors are not required in order for amitriptyline to cause antinociception in mice, but they are required to see caffeine reversal of this antinociceptive effect. When A1Rs are present, actions of amitriptyline may, however, partly depend on A1Rs.     

The effects of triiodothyronine (T3) on heart rate,temperature and ECG measured with telemetry in freely moving mice

We have set up a telemetry system and recorded heart rate, electrocardiogram (ECG), body temperature and the locomotor activity in awake freely moving mice. The telemetry system (DATA Sciences, St Paul, MN, USA) comprises a transmitter implanted in the peritoneal cavity and a receiver (RA1010) placed underneath the home cage. The signal from the transmitter includes the electrical activity of the heart and the body temperature. The results show that four days after the surgical procedure the mice have recovered and regained a clear circadian rhythm. The heart rate varied under baseline conditions between 432 and 618 beats min^?1 and the body temperature between 35.1 and 37.7 °C (based on 60 min mean values). A clear time correlation between heart rate, body temperature and locomotor activity was found. As an evaluation of the method we injected T₃ s.c. during a period of 4 days. Further, we were interested in whether it was possible to measure an integrated physiological response to T₃ and further investigate the time course for the effect. After one day of treatment with triiodothyronine there was a significant increase in body temperature and locomotor activity. The increase in heart rate was seen after 2 days. The ECG recording revealed a significantly shortened QT_end- and QRS-time. No significant difference in the PQ-time was found. This method may be of great importance in studies of genetically manipulated mice.     

The stimulatory effects of caffeine with oseltamivir (Tamiflu) on light–dark behavior and open-field behavior in mice

Abnormal behaviors and death associated with the use of oseltamivir (Tamiflu^®) have emerged as a major issue in influenza patients taking the drug. Here, we investigated the mechanisms underlying the effects of oseltamivir on the behavior of mice using light–dark and open-field preference tests. Oseltamivir (75 and 150 mg/kg, intraperitoneally (i.p.)) alone affected neither time spent in the open area in the light–dark preference test nor ambulation in the open-field test at 2 h post-injection. However, a non-selective adenosine A₁/A₂ receptor antagonist, caffeine (10 mg/kg, i.p.) in combination with oseltamivir (150 mg/kg, i.p.) increased time spent in the open area in the light–dark preference test. This enhancement was not inhibited by a benzodiazepine receptor antagonist, flumazenil (10–20 mg/kg, subcutaneously (s.c.)). Enhancement of ambulation in the open-field test was also observed when caffeine (10 mg/kg, i.p.) was combined with oseltamivir (150 mg/kg, i.p.). This enhancement was inhibited by a dopamine D₂ receptor antagonist, haloperidol (0.1 mg/kg, s.c.). Furthermore, an adenosine A₂ receptor antagonist, SCH58261 (3 mg/kg, i.p.) in combination with oseltamivir (150 mg/kg, i.p.) increased ambulation in the open-field test, while an adenosine A₁ receptor antagonist, DPCPX (1–3 mg/kg, i.p.) did not. These findings suggest that the actions of oseltamivir may involve the dopamine and adenosine systems. Our findings suggest that due to the interaction between central blockade of adenosine A₂ receptors by caffeine, and oseltamivir-induced behavioral changes, patients being treated with oseltamivir should be closely monitored.     

Modulation of morphine-induced antinociception in mice by histamine H3-receptor ligands

The effects of compounds active at histamine H₃-receptors on morphine-induced antinociception have been investigated in thermal and chemical tests in mice; tail-immersion (50°C) and hot-plate (49° and 55°C) tests and acetic acid-induced writhing. Neither (R)α-methylhistamine, a specific agonist, (S)α-methylhistamine, a chemical control, nor thioperamide, an antagonist, had any antinociceptive action alone but thioperamide (3 mg kg⁻¹) attenuated the effects of morphine in the tailimmersion test wile (R)α-methylhistamine (1 mg kg⁻¹), but not the (S) isomer, potentiated its effects in the hot-plate test at 55°C. These results are consistent with the morphine potentiation seen with H₁-antagonists and suggest that central histaminergic mechanisms can modulate opioid actions.

     

Effect of different xanthines and phosphodiesterase inhibitors on c-fos expression in rat striatum

It has previously been shown that caffeine, in a dose-dependent manner, increases the expression of the protooncogene c-fos in the rat brain, predominantly in the caudate-putamen and tuberculum olfactorium. In this study we examined the effect of related xanthines and of selective phosphodiesterase inhibitors on c-fos expression. The effect of caffeine (75 mg kg¹) was mimicked by 3-isobutyl-l-methyl xanthine (IBMX) (25 mg kg^-1) and theophylline (100 mg kg^-1) but not by 8-p-sulfophenyltheophylline (10mgkg¹), enprofylline, theobromine or paraxanthine (each at 100mgkg¹). Moreover, the cyclic AMP-selective phosphodiesterase inhibitors rolipram (10 or 20mgkg^_1) and SQ 20006 (25 mg kg ^:) and the cyclic GMP-selective phosphodiesterase inhibitor zaprinast (10 mg kg^-1) failed to induce c-fos in striatum, but caused a clear-cut induction in the overlying cerebral cortex. Thus, c-fos is induced in rat striatum following adminstration of caffeine and other xanthines that (provided they enter the brain) block adenosine receptors, suggesting an involvement of central adenosine receptors. Inhibition of cyclic nucleotide phosphodiesterase does not appear to play any important role in c-fos induction by the xanthines.     

Caffeine reverses antinociception by oxcarbazepine by inhibition of adenosine A1 receptors: Insights using knockout mice

Oxcarbazepine is an anticonvulsant drug that has been explored as a novel therapeutic agent to treat neuropathic pain in humans. It produces antinociception in several preclinical models of pain, and these actions are blocked by methylxanthine adenosine receptor antagonists which implicates adenosine it its actions. In this study, the antinociceptive effect of oxcarbazepine, and the ability of caffeine to reverse its actions, were examined using the formalin test (2%) in wild-type mice and in mice lacking adenosine A₁ receptors by way of further exploring the involvement of adenosine in its actions. Oxcarbazepine produced dose-related suppression of formalin-evoked flinching responses in wild-type mice following both systemic and intraplantar administration, and this action was reversed by systemic and intraplantar administration of caffeine, respectively. The ability of oxcarbazepine to inhibit flinching after systemic and intraplantar administration was unaltered in homozygous (−/−) and heterozygous (+/−) adenosine A₁ receptor knockout mice. However, caffeine no longer reversed this antinociception. Our results indicate that, while adenosine A₁ receptors are not required for oxcarbazepine to produce antinociception in knockout mice, such receptors are essential in order to see caffeine reversal of this antinociceptive effect.     

Adenosine A1 receptors and vascular reactivity

Aim: Blood pressure is higher in A₁ receptor knock-out (A₁R−/−) mice than in wild type litter mates (A₁R+/+) and we have examined if this could be related to altered vascular functions. Methods: Contraction of aortic rings and mesenteric arteries were examined. To examine if the adenosine A₁ receptor-mediated contraction of aortic muscle was functionally important we examined pulse pressure (PP) and augmentation index (AIX) using a sensor that allows measurements of rapid pressure transients. Results: Contraction of aortic rings to phenylephrine and relaxation to acetylcholine were similar between genotypes. The non-selective adenosine receptor agonist N-ethyl carboxamido adenosine (NECA) enhanced the contractile response, and this was eliminated in aortas from A₁R−/− mice. However, in mesenteric arteries no contractile response was seen and adenosine-mediated relaxation was identical between studied genotypes. A_2B adenosine receptors, rather than A_2A receptors, may be mainly responsible for the vasorelaxation induced by adenosine analogues in the examined mouse vessels. PP was higher in A₁R−/− mice, but variability was unaltered. AIX was not different between genotypes, but the NECA-induced fall was larger in A₁R−/− mice. Conclusions: The role of adenosine A₁ receptors in regulating vessel tone differs between blood vessels. Furthermore, contractile effects on isolated vessels cannot explain the blood pressure in A₁ knock-out mice. The A₁ receptor modulation of blood pressure is therefore mainly related to extravascular factors.     

Effects of Paraxanthine and Caffeine on Sleep,Locomotor Activity,and Body Temperature in Orexin/Ataxin-3 Transgenic Narcoleptic Mice

Study Objective:

Caffeine, an adenosine A1 and A2a receptor antagonist, is a widely consumed stimulant and also used for the treatment of hypersomnia; however, the wake-promoting potency of caffeine is often not strong enough, and high doses may induce side effects. Caffeine is metabolized to paraxanthine, theobromine, and theophylline. Paraxanthine is a central nervous stimulant and exhibits higher potency at A1 and A2 receptors, but has lower toxicity and lesser anxiogenic effects than caffeine.

Design:

We evaluated the wake-promoting efficacy of paraxanthine, caffeine, and a reference wake-promoting compound, modafinil, in a mice model of narcolepsy, a prototypical disease model of hypersomnia. Orexin/ataxin-3 transgenic (TG) and wild-type (WT) mice were subjected to oral administration (at ZT 2 and ZT14) of 3 doses of paraxanthine, caffeine, modafinil, or vehicle.

Results:

Paraxanthine, caffeine, and modafinil significantly promoted wakefulness in both WT and narcoleptic TG mice and proportionally reduced NREM and REM sleep in both genotypes. The wake-promoting potency of 100 mg/kg p.o. of paraxanthine during the light period administration roughly corresponds to that of 200 mg/kg p.o. of modafinil. The wake-promoting potency of paraxanthine is greater and longer lasting than that of the equimolar concentration of caffeine, when the drugs were administered during the light period. The wake-promotion by paraxanthine, caffeine, and modafinil are associated with an increase in locomotor activity and body temperature. However, the higher doses of caffeine and modafinil, but not paraxanthine, induced hypothermia and reduced locomotor activity, thereby confirming the lower toxicity of paraxanthine. Behavioral evaluations of anxiety levels in WT mice revealed that paraxanthine induced less anxiety than caffeine did.

Conclusions:

Because it is also reported to provide neuroprotection, paraxanthine may be a better wake-promoting agent for hypersomnia associated with neurodegenerative diseases.

Citation:

Okuro M; Fujiki N; Kotorii N; Ishimaru Y; Sokoloff P; Nishino S. Effects of paraxanthine and caffeine on sleep, locomotor activity, and body temperature in orexin/ataxin-3 transgenic narcoleptic mice. SLEEP 2010;33(7):930-942.     

Ergogenic effect of varied doses of coffee-caffeine on maximal aerobic power of young African subjects

Background

Caffeine one of the readily available stimulants consumed daily by more than 80% of the world''s population, making it the most widely consumed drug in history. The objective of this study was to determine the effects of different doses (5, 10 &15 mg.kg⁻¹) of caffeine per kilogram body weight on maximal aerobic power of normal young black African (Nigerian) male adults.

Method

Twenty apparently healthy young male adults volunteers, participated. A repeated measure four randomized crossover (counter balanced) double blind design was used in data collection. Subjects engaged in 20 meter shuttle run test (20 MST) one hour post caffeine (5, 10 & 15 mg.kg⁻¹) and placebo doses ingestion. Endurance performance index (VO₂ max, run time & number of exercise laps) were measured and recorded.

Result

Repeated measures ANOVA was used to assess the level of significant difference between caffeine doses and placebo dose in VO₂ max, run time and number of exercise laps. The result showed no significant effect of caffeine doses over placebo dose.

Conclusion

It was concluded that caffeine dose of up to 15mg/kg seems not to have any ergogenic effect on maximum aerobic power of young black African male adults.     

In silico study of naphtha [1, 2-d] thiazol-2-amine with adenosine A2A receptor and its role in antagonism of haloperidol-induced motor impairments in mice

Loss of dopaminergic nigrostriatal neurons in the substantia nigra leads to Parkinson's disease (PD). Adenosine A_2A receptors (A_2ARs) have been anticipated as novel therapeutic target for PD. A_2ARs potentiate locomotor behavior and are predominantly expressed in striatum. Naphtha [1, 2-d] thiazol-2-amine (NATA), a tricyclic thiazole have been studied as new anti-Parkinsonian compound. AutoDock analysis and pharmacophore study of NATA with known A_2AR antagonists explicit its efficacy as a possible adenosine receptor antagonist. In vivo pharmacology of NATA showed reduction of haloperidol (HAL)-induced motor impairments in Swiss albino male mice. Relatively elevated levels of dopamine in NATA pre-treated mice are suggestive of its possible role as neuromodulator in PD.     

Fat content affects heat capacity: a study in mice

The specific heat capacity of the human body is a widely used number in calculations related to studies in energy metabolism and temperature regulation. A value of 3.47 kJ kg^-1 K^-1 has been used during the last century, but there have been no direct studies on the dependence of this value on the body fat content. We present here results of measurements of the specific heat capacity of lean and obese mice. The mice were killed and heated to 40.6 d?C, transferred to a calorimeter and the specific heat capacity was determined from body mass of the mice and temperature rise of water in the calorimeter. We measured the fat fraction of body mass (m_t), fat free body mass and water content of the mice and found a marked dependency of the specific heat capacity (C_p) on body fat. In obese mice (fat content 52.76% body wt) the heat capacity was 2.65 kJ kg^-1 K^-1 and in lean mice (fat content 7.55% body wt) the heat capacity was 3.66 kJ kg^-1 K^-1     

Maternal glutamate intake during gestation and lactation regulates adenosine A1 and A2A receptors in rat brain from mothers and neonates

Pregnant rats were treated daily with 1 g/L of l-glutamate in their drinking water during pregnancy and/or lactation. The effect on adenosine A₁ receptor (A₁R) and A_2A receptor (A_2AR) in brains from both mothers and 15-day-old neonates was assayed using radioligand binding and real time PCR assays. Mothers receiving l-glutamate during gestation, lactation, and throughout gestation and lactation showed a significant decrease in total A₁R number (water+water, 302±49 fmol/mg; l-glutamate+water, 109±11 fmol/mg, P<0.01; water+l-glutamate, 52±13 fmol/mg, P<0.01; l-glutamate+l-glutamate, 128±33 fmol/mg, P<0.05). No variations were detected in the Kd parameter. Concerning adenosine A_2AR, radioligand binding assays revealed that Bmax parameter remains unaltered in maternal brain in response to glutamate exposure. However, Kd parameter was significantly decreased in all l-glutamate-treated groups (water+water, 5.3±1.3 nM; l-glutamate±water, 0.5±0.1 nM; water+l-glutamate, 0.9±0.1 nM; l-glutamate±l-glutamate, 0.7±0.1 nM, P<0.01 in all cases). In both male and female neonates, A₁R was also decreased after long-term glutamate exposure during gestation, lactation, and gestation plus lactation (male neonates: water+water, 564±68 fmol/mg; l-glutamate+water, 61±8 fmol/mg; water+l-glutamate, 95±20 fmol/mg; l-glutamate+l-glutamate, 111±15 fmol/mg; P<0.01 in all cases; female neonates: water+water, 216±35 fmol/mg; l-glutamate+water, 59±9 fmol/mg; water+l-glutamate, 139±16 fmol/mg; l-glutamate+l-glutamate, 97±14 fmol/mg; P<0.01 in all cases). No variations were found in mRNA level coding adenosine A₁R in maternal or neonatal brain. Concerning adenosine A_2AR, radioligand binding assays revealed that Bmax parameter was significantly increased in male and female neonates exposed to l-glutamate during lactation (male neonates: water+water, 214±23 fmol/mg; water+l-glutamate, 581±49 fmol/mg; P<0.01; female neonates: water+water, 51±10 fmol/mg; water+l-glutamate, 282±52 fmol/mg; P<0.05). No variations were found in mRNA level coding adenosine A_2AR in maternal or neonatal brain. In summary, long-term l-glutamate treatment during gestation and lactation promotes a significant down-regulation of A₁R in whole brain from both mother and neonates and a significant up-regulation of A_2AR in neonates exposed to l-glutamate during lactation.     

Effects of aerobic fitness on hypohydration-induced physiological strain and exercise impairment

Aim: Hypohydration exacerbates cardiovascular and thermal strain and can impair exercise capacity in temperate and warm conditions. Yet, athletes often dehydrate in exercise, are hypervolaemic and have less cardiovascular sensitivity to acute hypervolaemia. We tested the hypothesis that trained individuals have less cardiovascular, thermoregulatory and performance affect of hypohydration during exercise. Methods: After familiarization, six trained [O_{2 peak} = 64 (SD 8) mL kg⁻¹ min⁻¹] and six untrained [O_{2 peak} = 45 (4) mL kg⁻¹ min⁻¹] males cycled 40 min at 70%O_{2 peak} while euhydrated or hypohydrated by 1.5–2.0% body mass (crossover design), before a 40-min work trial with euhydration or ad libitum drinking (in Hypohydration trial), in temperate conditions (24.3 °C, RH 50%, v_a = 4.5 m s⁻¹). Baseline hydration was by complete or partial rehydration from exercise+heat stress the previous evening. Results: During constant workload, heart rate and its drift were increased in Hypohydration compared with Euhydration for Untrained [drift: 33 (11) vs. 24 beats min⁻¹ h⁻¹ (10), 95% CI 5–11] but not Trained [14 (3) vs. 13 beats min⁻¹ h⁻¹ (3), CI −2 to 3; P = 0.01 vs. Untrained]. Similarly, rectal temperature drift was faster in Hypohydration for Untrained only [by 0.57 °C h⁻¹ (0.25); P = 0.03 vs. Trained], concomitant with their reduced sweat rate (P = 0.05) and its relation to plasma osmolality (P = 0.03). Performance power tended to be reduced for Untrained (−13%, CI −35 to 2) and Trained (−7%, CI: −16 to 1), without an effect of fitness (P = 0.38). Conclusion: Mild hypohydration exacerbated cardiovascular and thermoregulatory strain and tended to impair endurance performance, but aerobic fitness attenuated the physiological effects.     

Additive hypothermic effects of the 5‐HT1A receptor agonist 8‐OH‐DPAT and the dopamine D2/3 receptor agonist 7‐OH‐DPAT in the rat

The objective of this study was to examine possible interactions between serotonergic and dopaminergic agents lowering core temperature via stimulation of 5‐HT_1A and dopamine (DA) D₂ receptors, respectively. The effects of the 5‐HT_1A receptor agonist (±)‐8‐hydroxy‐2‐(di‐n‐propylamino)tetralin HBr (8‐OH‐DPAT) and the DA D_2/3 receptor agonist 7‐OH‐DPAT on core temperature was monitored in adult male Wistar rats, approximately 300 g body weight. The temperature probe was connected to a PC‐assisted temperature instrument, and an automated printer device was activated when the temperature reading had stabilized (±0.1 °C) for 10 s. As expected, 7‐OH‐DPAT [0.5 and 2.0 μmol kg^–1 subcutaneous (s.c.)] as well as 8‐OH‐DPAT (0.15–2.4 μmol kg^–1 s.c.), produced a dose‐dependent hypothermia. When combined, there were additive effects of the two compounds, although the effects of 7‐OH‐DPAT were attenuated by 8‐OH‐DPAT at the higher doses (0.6–2.4 μmol kg^–1), in all probability because of emerging DA D₂ receptor blocking properties of the latter compound.     

Effects of caffeine ingestion on endurance racing in heat and humidity

A hot and humid environment can be detrimental to race performance. Caffeine, on the other hand, has been shown to be an ergogenic aid for improving endurance performance. To examine the influence of caffeine ingestion on race performance during high heat stress, seven endurance trained competitive road racers aged between 23 and 51 years (five men, two women) performed three maximal effort 21-km road races outdoors in hot and humid conditions. The caffeine dose, randomly assigned in a double-blind fashion, consisted of either 0, 5, or 9 mg · kg⁻¹ body mass. During each run, the subjects were allowed to drink waterad libitum at each 5-km point. Blood samples were obtained immediately before and after each run and analysed for changes in concentrations of Na⁺, K⁺, glucose, lactate, and hematocrit. Pre and postrun data were also collected for body mass and tympanic membrane temperature. Race times were not significantly different among the races or caffeine doses, with the average times within 1.1% of each other. In addition, none of the other variables measured varied significantly among the races or caffeine doses. In summary, caffeine intake did not affect race performance. Therefore it was concluded from our study that caffeine is not of ergogenic benefit in endurance races during high heat stress.     

Longitudinal bone growth is impaired by direct involvement of caffeine with chondrocyte differentiation in the growth plate

We showed previously that caffeine adversely affects longitudinal bone growth and disrupts the histomorphometry of the growth plate during the pubertal growth spurt. However, little attention has been paid to the direct effects of caffeine on chondrocytes. Here, we investigated the direct effects of caffeine on chondrocytes of the growth plate in vivo and in vitro using a rapidly growing young rat model, and determined whether they were related to the adenosine receptor signaling pathway. A total of 15 male rats (21 days old) were divided randomly into three groups: a control group and two groups fed caffeine via gavage with 120 and 180 mg kg⁻¹ day⁻¹ for 4 weeks. After sacrifice, the tibia processed for the analysis of the long bone growth and proliferation of chondrocytes using tetracycline and BrdU incorporation, respectively. Caffeine‐fed animals showed decreases in matrix mineralization and proliferation rate of growth plate chondrocytes compared with the control. To evaluate whether caffeine directly affects chondrocyte proliferation and chondrogenic differentiation, primary rat chondrocytes were isolated from the growth plates and cultured in either the presence or absence of caffeine at concentrations of 0.1–1 mm , followed by determination of the cellular proliferation or expression profiles of cellular differentiation markers. Caffeine caused significant decreases in extracellular matrix production, mineralization, and alkaline phosphatase activity, accompanied with decreases in gene expression of the cartilage‐specific matrix proteins such as aggrecan, type II collagen and type X. Our results clearly demonstrate that caffeine is capable of interfering with cartilage induction by directly inhibiting the synthetic activity and orderly expression of marker genes relevant to chondrocyte maturation. In addition, we found that the adenosine type 1 receptor signaling pathway may be partly involved in the detrimental effects of caffeine on chondrogenic differentiation, specifically matrix production and mineralization, as evidenced by attenuation of the inhibitory effects of caffeine by blockade of this receptor. Thus, our study provides novel information on the integration of caffeine and adenosine receptor signaling during chondrocyte maturation, extending our understanding of the effect of caffeine at a cellular level on chondrocytes of the growth plate.