beta -Adrenergic receptor number in human lymphocytes is inversely correlated with aerobic capacity

Nobuharu Fujii1, Sachiko Homma2, Fumio Yamazaki3, Ryoko Sone4, Takeshi Shibata1, Haruo Ikegami5, Kazuo Murakami1, and Hitoshi Miyazaki1

1 Gene Experiment Center, Institute of Applied Biochemistry, University of Tsukuba, Tsukuba-City 305; 2 Research Institute of Physical Fitness, Japan Women's College of Physical Education, Tokyo 157; 3 School of Health Science, University of Occupational and Environmental Health, Kitakyushu 807; 4 Department of Exercise and Health Science, Faculty of Education, University of Yamaguchi, Yamaguchi 753; and 5 Department of Physical Education, International Budo University, Katsuura 299-52, Japan

    ABSTRACT
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Abstract
Introduction
Methods
Results
Discussion
References

In the present study, the relationships between beta -adrenergic receptor (beta -AR) expression and aerobic capacity evaluated by maximal oxygen consumption (<A><AC>V</AC><AC>˙</AC></A><SC>o</SC><SUB>2 max</SUB>) and oxygen consumption level at ventilatory threshold (VO2@VT) were investigated. Seventeen physically untrained and 25 trained men participated in the study. After supine resting, the peripheral blood was sampled for preparation of lymphocytes, the model cell used to analyze the beta -AR state. The total number of beta -AR in lymphocytes (beta -ARtotal) was inversely correlated with the VO2 max (r = -0.368; P < 0.05) and the VO2@VT (r = -0.359; P < 0.05). Similar relationships were also observed between the number of beta -AR in cell surface and both VO2 max (r = -0.491; P < 0.05) and VO2@VT (r = -0.498; P < 0.05). However, no correlation was obtained between the number of beta -AR in intracellular compartments and either VO2 max or VO2@VT. The beta 2-AR mRNA level quantified by the use of competitive reverse transcription-polymerase chain reaction was inversely correlated with VO2@VT (r = -0.567; P < 0.05) and positively correlated with beta -ARtotal (r = 0.521; P < 0.05). These findings suggest that the beta -AR number in lymphocytes is inversely correlated with aerobic capacity. This relationship may be explained by downregulation of beta -AR, including internalization with subsequent degradation of the receptors and inhibition of the beta -AR biosynthesis.

catecholamine; downregulation; competitive reverse transcription-polymerase chain reaction

    INTRODUCTION
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Abstract
Introduction
Methods
Results
Discussion
References

PHYSICAL TRAINING is associated with cardiovascular adaptations such as bradycardia at rest (33) and lower heart rate and blood pressure responses during submaximal exercise (7). These phenomena may be explained at least in part by a decrease in the number of myocardial beta -adrenergic receptors (beta -AR) causing reduced sympathomimetic effects (30). In in vitro experiments, prolonged agonist exposure induced the loss of beta -AR at the cell surface (35). In addition, 24-h integrated plasma catecholamine concentrations are greater in physically trained men than in untrained men (9). Therefore, physical training seems to induce the loss of beta -AR and the attenuation of cellular responsiveness to sympathoadrenomedullary activity.

The number of beta -AR, the responsiveness of beta -AR to its agonist, and the content of stimulatory guanine nucleotide-binding protein in human lymphocytes are correlated with those in the myocardium (4, 16). Consequently, in humans, the effects of physical training on beta -AR have been studied using lymphocytes derived from peripheral blood as a model cell (22). However, so far, conflicting results have been obtained. Butler et al. (5) and Ohman et al. (29) found a significant reduction in the beta -AR number induced by training, but, conversely, several other investigators observed increased beta -AR number in trained subjects (20, 23). Moreover, Frey et al. (10) failed to demonstrate any relation between beta -AR number and physical fitness. In this regard, Jost et al. (17) reported that long-distance runners and swimmers had fewer beta -AR during the endurance training period (i.e., relatively aerobic exercise training) compared with sedentary individuals, but slightly more beta -AR were seen during the high-intensity training period (i.e., relatively anaerobic exercise training). Their report indicates a possibility that the controversial results acquired in previous studies may have arisen due to the difference in the exercise training conditions or the change in physiological functions such as the aerobic energy system after training variation. Thus different conclusions would be obtained by various research groups, depending on the variation of physical training such as endurance training and high-intensity training, even though the same types of athletes might have constituted the subjects of the studies. If so, group comparison would not be a suitable design for this kind of study. Therefore, it is possible that some relations between the physical training and the beta -AR number would be detected by characterizing the aerobic capacity in individual subjects.

The present study was thus conducted to clarify the effect of physical training on the beta -AR number in human lymphocytes by examining this effect as a function of aerobic capacity evaluated by maximal oxygen consumption (VO2 max) and oxygen consumption level at ventilatory threshold (VO2@VT). To obtain more detailed information on the underlying mechanism of the change in the beta -AR number with aerobic capacity, we examined the number of beta -AR in both the cell surface (beta -ARsurface) and intracellular compartment (beta -ARintra) in intact lymphocytes by measuring the binding of the lipophilic radioligand [125I]iodocyanopindolol ([125I]ICYP) in the presence and absence of the hydrophilic or lipophilic nonradioactive antagonist. Moreover, we used the competitive reverse transcription (RT)-polymerase chain reaction (PCR) method, which is effective in quantifying the mRNA level in small samples such as the lymphocytes in blood derived from human subjects, to elucidate the effects of physical training on the beta -AR mRNA level.

    METHODS
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Abstract
Introduction
Methods
Results
Discussion
References

Subjects. Seventeen physically untrained men with age range of 20-27 yr and 25 physically trained men with age range of 18-22 yr participated in this study after having given written informed consent. The trained subjects were collegiate long-distance runners (n = 13), short-distance runners (n = 6), and baseball players (n = 6). The untrained men were sedentary or participated only in leisure sports. None of them was receiving any medication, and a thorough clinical examination failed to demonstrate any abnormalities in their health.

Experimental protocol. An indwelling catheter with a three-way stopcock was inserted into a forearm vein. After the subject lay supine for 20 min in a quiet room, a 40-ml blood sample was withdrawn for preparation of lymphocytes and determination of plasma catecholamine concentrations. The total number of beta -AR in lymphocytes (beta -ARtotal) and plasma catecholamine concentrations were quantified in all subjects, except that in one subject the plasma catecholamine concentrations could not be measured because of insufficient blood sampling. The beta -ARsurface and beta -ARintra were analyzed in 24 subjects (9 untrained men, 7 long-distance runners, 4 short-distance runners, and 4 baseball players), and the beta 2-AR mRNA level was assayed in 18 subjects (8 untrained men, 6 long-distance runners, 2 short-distance runners, and 2 baseball players) because of limitation on the amount of blood that could be obtained from a subject. For measurement of beta -ARtotal, beta -ARsurface and beta -ARintra, beta 2-AR mRNA level, and plasma catecholamine concentrations, 24, 12, 12, and 4 ml of blood were used, respectively. After blood was sampled, the subject performed bicycle ergometry in a sitting position until exhaustion. Every 2 min, the workload was increased by 20 watts in untrained and 25 watts in trained subjects. Respiratory gas analysis was performed with an Oxycon-4 (Mijnhard) and was used to determine minute ventilation (VE), minute oxygen consumption (VO2), and minute carbon dioxide production (VCO2). These parameters were averaged over the last 30 s in each stage and supplied for the determination of VO2 max and VO2@VT.

Determination of VO2 max and VO2@VT. VO2 max was determined as the VO2 averaged over the last 30 s in the final stage. Ventilatory threshold was determined by visual inspection after consideration of events during incremental exercise, as a deviation of the incremental rate of VE plotted against VO2 from the linear increase, and an increase of VE/VO2 plotted against VO2 without an increase of VE/VCO2 (8).

Measurement of the lymphocyte beta -AR number. Lymphocytes were prepared by the method of Bøyum (2). Whole blood was diluted with an equal volume of phosphate-buffered saline (PBS), and an aliquot (17 ml) was then carefully layered on 12 ml of Ficoll-Paque (Pharmacia). The tubes were centrifuged at 400 g for 40 min. After removal of the plasma, the lymphocytes were harvested carefully, washed three times with 10 vol of PBS, and then resuspended in 10 mM Tris, 154 mM NaCl, and 0.55 mM ascorbic acid, pH 7.4 (buffer A). For binding assay, intact lymphocytes (1 × 106 cells) were incubated with various concentrations (1-180 pM) of [125I]ICYP in 0.5 ml of buffer A containing 0.05% bovine serum albumin (BSA). Propranolol (2 µM) and CGP-12177A (2 µM) were used to detect nonspecific binding (26). The difference in [125I]ICYP binding with and without the hydrophobic ligand propranolol represents the total specific binding to beta -ARtotal. The difference in binding with and without the hydrophilic ligand CGP-12177A (Ciba-Geigy) indicates the specific binding to beta -ARsurface. The specific binding to beta -ARintra was obtained from the difference between the binding to beta -ARtotal and to beta -ARsurface. The incubation was carried out at 37°C for 40 min and stopped by placing the tubes on ice and adding 2 ml of ice-cold PBS containing 0.1% BSA to each tube. The samples were filtered through Whatman GF/D filters, and each filter was washed two times with an additional 3 ml of PBS. The radioactivity of the filters was determined in a gamma counter (ARC1000M; Aloka). The binding data were analyzed according to the method of Scatchard (32).

Isolation of total RNA. Total cellular RNA was isolated by Isogen (Nippon Gene) containing guanidinium isothiocyanate and phenol (6). Briefly, aliquots of the cells were lysed by pipetting in a mixture containing Isogen and chloroform. After centrifugation at 4°C, the aqueous phase was precipitated in isopropanol. The RNA pellet was washed one time in 70% ethanol, dried, and dissolved in diethylpyrocarbonate-treated water. Genomic DNA was removed by digestion with RNase-free DNase I (Nippon Gene) for 1 h at 37°C. RNA was checked by 1% agarose gel electrophoresis in the presence of 0.66 M formaldehyde. The purity of RNA was checked spectrophotometrically by the ratio of 260 nm to 280 nm and electrophoretically by the presence of intense bands of 18S and 28S RNA. The yield was determined spectrophotometrically at 260 nm.

Preparation of a deletion-mutated beta -AR cDNA fragment for competitive RT-PCR analysis. A 401-bp fragment containing part of the coding region of the human beta 2-adrenergic receptor (beta 2-AR) gene (73-473; nucleotides numbered sequentially from the translation initiation site) was amplified with the following primers: 5'-ACGCAGCAAAGGGACGAG-3' (5'-sense primer) and 5'-CACACCATCAGAATGATCAC-3' (3'-antisense primer; see Ref. 11). These fragments were phosphorylated at their 5'-ends and inserted into the EcoR V site of pBluescript KS(+) (Stratagene). The resultant plasmid was digested with Msc I and BstE II, blunt-ended with T4 DNA polymerase, and self-ligated to generate a plasmid containing the insert lacking the Msc I-BstE II fragment (62 bp). This plasmid was amplified by PCR using the primers described above, and the resultant deletion-mutated beta 2-AR cDNA (339 bp) was resolved by polyacrylamide gel electrophoresis. After recovery, the cDNA fragment was quantified and used as a competitor for competitive RT-PCR.

Competitive RT-PCR. Total RNA (1.8 µg) was reverse transcribed to cDNA in the presence of 10 U/µl cloned Moloney murine leukemia virus reverse transcriptase (GIBCO-BRL), 125 µM each of dNTPs, 10 mM dithiothreitol, 2 U/µl ribonuclease inhibitor (Takara), 5 ng/µl pd(N)6 random primers, and RT buffer (50 mM Tris · HCl, pH 8.3, 75 mM KCl, and 3 mM MgCl2) to a total volume of 20 µl. A corresponding aliquot of cDNA mixture synthesized from 100 ng of total RNA was subjected to competitive PCR analysis using the same primers (1 µM each) described above in the presence of various amounts of the competitors to quantify the PCR products. Reaction solution for PCR consisted of 10 mM Tris · HCl, pH 8.3, 50 mM KCl, 1.5 mM MgCl2, 0.01% (wt/vol) gelatin, 125 µM each of dNTPs, 0.01 µCi/µl [alpha -32P]dCTP, and 0.02 U/µl Amplitaq polymerase (Perkin-Elmer). PCR was performed by 28 cycles of denaturation at 94°C for 1 min, annealing at 56°C for 1 min, and extension at 72°C for 1.5 min. Thereafter, the incubation was continued at 72°C for 8.5 min to complete the polymerization. The PCR products were size-fractionated on 5% acrylamide gels. The gels were dried and analyzed using a computer-based imaging system (BAS 2000; Fuji). The amount of beta 2-AR mRNA was then calculated by extrapolating from the intersection of the curves, where the amounts of target and competitor were equivalent [log(target/competitor) = 0] to the x-axis (12). Resultant values of beta 2-AR mRNA were normalized with the RT-PCR products of glyceraldehyde-6-phosphate dehydrogenase mRNA quantified in an independent experiment series (12).

Determination of beta -AR subtypes expressed in human lymphocytes by RT-PCR. To determine the beta -AR subtypes expressed in lymphocytes, RT-PCR was carried out under the same conditions as described above except that the competitor was not added, and denaturing, annealing, and extension reactions proceeded 30 times at 94°C for 1 min, 50°C for 1 min, and 72°C for 1.5 min, respectively. The following PCR oligonucleotide primers that hybridize to the conserved sequence among the human beta 1-adrenergic receptor (beta 1-AR), beta 2-AR, and beta 3-adrenergic receptor (beta 3-AR) cDNAs were used: sense 5'-GTCTCCTTCTACGTTCC-3' and antisense 5'-AAGAAGGGCATCCAGCAGAG-3' (Fig. 1A). Amplified cDNAs of beta 1-, beta 2-, and beta 3-ARs should give 337-, 254-, and 296-bp fragments, respectively. These products were electrophoretically separated on 4% acrylamide gels. The gels were dried and analyzed using a computer-based imaging system (BAS 2000; Fuji).


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Fig. 1.   Subtype distribution of beta -adrenergic receptors (beta -AR) in human lymphocytes. RT-PCR was performed by using the oligonucleotide primers that hybridize the conserved sequence among the human beta 1-, beta 2-, and beta 3-AR cDNA. A: nucleotide sequences of the human beta 1-, beta 2-, and beta 3-AR cDNA that are hybridized by PCR primers. Incomplementary sequences to PCR primers are boxed. Nucleotides are numbered sequentially from the translation initiation site. B: size fraction of beta -AR subtype cDNAs amplified from human lymphocyte RNA by RT-PCR. C: quantitative comparison of the mRNA expression level of beta -AR subtypes in lymphocytes.

Determination of plasma catecholamines. Plasma norepinephrine and epinephrine concentrations were quantified using high-performance liquid chromatography with electrochemical detection (13).

Statistics. Pearson's formula was used to calculate the correlation coefficients. Comparisons among groups were made by one-way ANOVA. Probability value of <0.05 was considered significant.

    RESULTS
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Methods
Results
Discussion
References

We first examined the subtype proportion of beta -AR in human lymphocytes by RT-PCR using oligonucleotide primers that hybridize to the conserved sequence among the human beta 1-, beta 2-, and beta 3-AR cDNAs (Fig. 1A). Because the melting temperature of these primers to the cDNA of each subtype is identical, radioactivity of resultant PCR products that are size-fractionated by electrophoresis indicates the mRNA level of each beta -AR subtype. As shown in Fig. 1, B and C, beta 2-AR mRNA level was predominant. In contrast, little amount of beta 1-AR mRNA (~1.5% of the total specific radioactivity) and no beta 3-AR mRNA were detected. Similar results were also obtained from separate PCR amplification of each beta -AR subtype using specific primers for them in different reaction tubes (data not shown). Therefore, predominant amplification of beta 2-AR mRNA was not artificially caused by the coamplification of all three subtype fragments in a single reaction tube. These results strongly demonstrate that almost every beta -AR is the beta 2-AR subtype in human lymphocytes.

As presented in Fig. 2, the beta -ARtotal was inversely correlated with VO2 max and VO2@VT. Similar relationships between the beta -ARsurface and both VO2 max and VO2@VT were also observed (Fig. 3). In contrast, no correlation was obtained between beta -ARintra and either VO2 max or VO2@VT (Fig. 4). The beta 2-AR mRNA level was inversely correlated with VO2@VT (Fig. 5) and tended to be correlated with VO2 max (P = 0.07). As indicated in Fig. 6, the beta -ARtotal was positively correlated with the beta 2-AR mRNA level.


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Fig. 2.   Scatterplots of the relation of the total number of beta -AR in lymphocytes (beta -ARtotal) in blood samples obtained at rest with maximal oxygen consumption (VO2 max; A) and oxygen consumption level at ventilatory threshold (VO2@VT; B). Lymphocytes were prepared from blood samples obtained at rest just before bicycle ergometry. open circle , Untrained men; x, baseball player; square , short-distance runner; black-triangle, long-distance runner.


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Fig. 3.   Scatterplots of the relation of the number of lymphocyte beta -AR in cell surfaces (beta -ARsurface) with VO2 max (A) and VO2@VT (B). Lymphocytes were prepared from blood samples obtained at rest just before bicycle ergometry. open circle , Untrained men; x, baseball player; square , short-distance runner; black-triangle, long-distance runner.


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Fig. 4.   Scatterplots of the relation of the number of lymphocyte beta -AR in intracellular compartments (beta -ARintra) with VO2 max (A) and VO2@VT (B). Lymphocytes were prepared from blood samples obtained at rest just before bicycle ergometry. open circle , Untrained men; x, baseball player; square , short-distance runner; black-triangle, long-distance runner.


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Fig. 5.   Scatterplots of the relation of the lymphocyte beta 2-AR mRNA level with VO2 max (A) and VO2@VT (B). Lymphocytes were prepared from blood samples obtained at rest just before bicycle ergometry. open circle , Untrained men; x, baseball player; square , short-distance runner; black-triangle, long-distance runner.


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Fig. 6.   Relationship between beta -ARtotal and beta 2-AR mRNA level. open circle , Untrained men; x, baseball player; square , short-distance runner; black-triangle, long-distance runner.

Table 1 shows the group comparison of the aerobic capacity and the number of beta -AR in lymphocytes. Significant differences were observed for VO2 max and VO2@VT among the groups by ANOVA. However, in contrast to the results of cross-sectional analysis (Figs. 2-5), no statistically significant differences for the measurements on the beta -AR number and on the beta -AR mRNA level were seen among the groups, except that beta -ARtotal tended to be different (P < 0.15).

                              
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Table 1.   Comparisons of the aerobic capacity and the beta -adrenergic receptor expression level in lymphocytes among the groups

The results of correlation analysis of the relationship of each plasma catecholamine concentration with beta -ARtotal, beta -ARsurface, beta -ARintra, and beta 2-AR mRNA level are summarized in Table 2. A significant inverse correlation between the plasma epinephrine concentration and the beta -ARsurface was found. Plasma epinephrine concentration tended to be correlated with beta -ARtotal (P = 0.07). A significant relationship between the plasma epinephrine concentration and the beta 2-AR mRNA level could not be demonstrated. The plasma norepinephrine concentration showed no correlation with any of the beta -ARtotal, beta -ARsurface, beta -ARintra, and beta 2-AR mRNA levels.

                              
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Table 2.   Correlation coefficients for plasma catecholamine concentrations with beta -adrenergic receptor number and mRNA level

No significant correlations were seen between VO2 max and both plasma epinephrine (r = 0.174) and plasma norepinephrine (r = 0.226) concentrations and between VO2@VT and both plasma epinephrine (r = 0.222) and plasma norepinephrine (r = 0.146) concentrations.

    DISCUSSION
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Introduction
Methods
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In the present study, lymphocyte beta -ARtotal was inversely correlated with VO2 max and VO2@VT. These results support the assertion by Butler et al. (5) and by Ohman et al. (29) that the beta -AR number in human lymphocytes is reduced by physical training. We also found that the inverse relationships were dependent on the change in the beta -ARsurface but not on that in the beta -ARintra. Furthermore, this is the first report indicating a significant inverse correlation between the beta 2-AR mRNA level and VO2@VT and a positive association between the beta 2-AR mRNA level and beta -ARtotal. These results suggest that the reduction of beta -AR number is involved with the clonic adaptation to aerobic exercise.

Continued exposure of cells to beta -adrenergic agonists in vitro leads to a decrease in the number of functional beta -AR in the plasma membrane in a manner indicating the loss of receptors from the cell surface (15). Downregulation and sequestration have been identified as two phenomena that contribute to the agonist-mediated decrease in receptor number from the cell surface (35). Downregulation has been hypothesized to involve accelerated receptor degradation, presumably in lysosomes, as well as decreased receptor biosynthesis. Sequestration has been proposed to result from internalization of receptors for reactivation and recycling of desensitized receptors from the cell surface to the intracellular compartment depot in which receptors are not degraded. Because there was a significant inverse relationship between the plasma epinephrine concentration and beta -ARsurface in this study, it is possible that plasma epinephrine promotes the internalization of beta -AR from the cell surface to the intracellular space. If this internalization occurred as an event of sequestration, beta -ARintra would increase and be positively correlated with plasma epinephrine. In this study, however, no significant correlation was found between the two, suggesting that plasma epinephrine may be associated with the degradation of beta -AR. Dela et al. (9) reported that the 24-h integrated plasma catecholamine concentrations were two times as high in physically trained as in untrained subjects. Therefore, our findings suggest that the increased epinephrine secretion induced by daily training is associated with the decrease in beta -ARsurface due to the internalization with subsequent degradation of beta -AR. Furthermore, the increase in the plasma epinephrine concentration, rather than plasma norepinephrine concentration, appears to be associated with this phenomenon.

The significant relationship between beta -ARtotal and beta 2-AR mRNA level observed in this study suggests that the suppression of biosynthesis of beta -AR associated with the reduction in mRNA level also contributes to the decrease in beta -ARtotal after physical training. It is known that chronic stimulation of cells with beta -adrenergic agonist reduces the steady-state level of beta 2-AR mRNA in vitro (14). However, in our in vivo study, the plasma catecholamine concentrations were not correlated with beta 2-AR mRNA level. Therefore, other factors may contribute to the reduction of the beta 2-AR mRNA level. It should be noted that the regulation of beta 2-AR mRNA level might be expected to be a reflection of the chronic effect of training and thus of the chronic change in the concentrations of catecholamines. The measurement of 24-h integrated plasma catecholamine concentrations as reported by Dela et al. (9), therefore, may be necessary for examining the effect of catecholamines on the beta 2-AR mRNA in vivo.

Jost et al. (17) reported that long-distance runners and swimmers had fewer beta -AR during the endurance training period compared with sedentary individuals, but slightly more beta -AR were seen during the high-intensity training period. Their finding indicates that the difference in the energy supply system recruited during exercise training may affect the expression of beta -AR. Thus we hypothesized that the different conclusions obtained by various research groups (5, 10, 20, 23, 29) may depend on the variation of physical training such as endurance training and high-intensity training, even though the same types of athletes might have constituted the subjects of the studies. In this study, no significant difference of the beta -AR number among the groups was obtained by group comparison analysis using ANOVA. In contrast, inverse relationships between the beta -AR number and the aerobic capacity were detected by examining the change in beta -AR number and mRNA level as a function of aerobic capacity in a heterogeneous group of subjects who had wide distribution of VO2 max and VO2@VT. These results may support our hypothesis.

Because there is a report (21) indicating that beta 2-AR couples to adenylyl cyclase with a greater efficacy than beta 1-AR, beta 2-AR may have an important role in regulating the cardiac function despite the lower expression level compared with beta 1-AR in human heart (3). Actually, overexpression of human beta 2-AR in heart of transgenic mice (25) exhibits functional changes more drastically than overexpression of human beta 1-AR (1). Therefore, if the regulation of the beta 2-AR content in lymphocytes reflects that in the heart, lymphocyte beta 2-AR analysis may provide significant information about the role of the beta -adrenergic system in the cardiac functions. Although we have no direct evidence on whether the change in the content of myocardial beta 2-AR as a result of exercise training parallels that in lymphocytes in this study, Michel et al. (24) reported that lymphocyte beta 2-AR number significantly correlates with the number of myocardial beta 2-AR under basal conditions.

Lymphocytes comprise distinct subsets that differ in the number of beta -AR (19). This feature raises the possibility that the change in composition of these subsets may be induced by physical training. Some reports suggest that chronic physical training induces no change of the lymphocyte subset distribution in human subjects (18, 27, 28). However, a few investigators have found changes in the percentages of the subsets associated with physical training, i.e., increased natural killer cell population in endurance-trained men (31) and increased T-suppressor/cytotoxic cell population and decreased T-helper cell population in distance runners (34). Our results might underestimate the effects of physical training on downregulation of beta -AR, since among lymphocytes the natural killer cell population has the most and the T-helper cell population the least beta -AR.

In conclusion, we found an inverse relationship between the beta -AR number in human lymphocytes and the aerobic capacity evaluated by VO2 max and VO2@VT. We consider that this inverse relationship may result from downregulation of the beta -AR, including internalization with subsequent degradation of the receptors and inhibition of the beta -AR biosynthesis after aerobic physical training.

    ACKNOWLEDGEMENTS

We are grateful to Dr. Yoshiharu Nabekura at the University of Tsukuba for managing the subjects. We thank Dr. Tetsuya Izawa at the Tokyo Metropolitan University for critical discussion. We appreciate the donation of CGP-12177A by K. G. Hofbauer and Dr. H. Kaufmann (Ciba-Geigy, Basel).

    FOOTNOTES

This study was supported in part by grants from the Meiji Life Foundation of Health and Welfare, Japan.

Address for reprint requests: H. Miyazaki, Gene Experiment Center, Univ. of Tsukuba, Tsukuba-City, Ibaraki 305, Japan.

Received 11 August 1997; accepted in final form 19 February 1998.

    REFERENCES
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Abstract
Introduction
Methods
Results
Discussion
References

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Am J Physiol Endocrinol Metab 274(6):E1106-E1112
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