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β‐Adrenoceptor subtypes in young and old rat ventricular myocytes: a combined patch‐clamp and binding study
Author(s) -
Cerbai E.,
Guerra L.,
Varani K.,
Barbieri M.,
Borea P.A.,
Mugelli A.
Publication year - 1995
Publication title -
british journal of pharmacology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.432
H-Index - 211
eISSN - 1476-5381
pISSN - 0007-1188
DOI - 10.1111/j.1476-5381.1995.tb16671.x
Subject(s) - isoprenaline , medicine , endocrinology , myocyte , patch clamp , stimulation , agonist , electrophysiology , antagonist , biology , chemistry , receptor
1 We used electrophysiological and binding techniques to assess the presence of β 1 and β 2 ‐adrenoceptors (β 1 AR and β 2 AR) in rat cardiac myocytes and to determine their ratio during aging. Experiments were performed in left ventricular myocytes enzymatically dissociated from the heart of 3‐(young) or 22‐month‐old (old) Wistar Kyoto rats 2 In patch‐clamp experiments, myocytes from old rats showed a prolonged action potential duration (at −20 mV: 41.7±3.6 vs 26.2±3.1 ms; at −60 mV: 154.4 ± 17.7 vs 87.1 ±6.9 ms, P < 0.05) and an augmented membrane capacitance (an index of cell size) (271.7±20.2 vs 164.3± 14.6 pF, P < 0.05) compared to young rats. β 2 AR stimulation, achieved by superfusing myocytes with the selective β 2 AR agonist, zinterol (10 μ m ) or with (−)‐isoprenaline (1 μ m ) in the presence of the selective β 1 AR antagonist, CGP 20712A (0.1 μ m ), significantly increased L‐type calcium current ( I ca,L ) in rat ventricular myocytes. The percentage increase was similar in both young and old rats, either with zinterol (26.9 ±3.6% and 24.2±2.8%, respectively) or isoprenaline plus CGP 20712A (30.4±3.7% and 22.4±4.1%, respectively). Isoprenaline alone (β 1 AR and β 2 AR stimulation) caused a much smaller increase in I Ca,L in old rats (58.4±12.1%) than in younger ones (95.3±8.1%) (P=0.067) 3 The number of βAR mg −1 protein, measured with saturation binding assays of the non selective βAR antagonist [ 3 H]‐CGP 12177 was 1989.4± 189.5 for 3‐ and of 1580.7± 161.5 for 22‐month‐old rats. Competition for [ 3 H]‐CGP 12177 binding by CGP 20712A gave biphasic curves which demonstrated two classes of binding sites. Densities (as percentages of total PAR density), and affinities for the two binding sites were: 80.4 ±2.2% ( K i = 6.6 ±1.3 nM) β 1 AR and 19.6 ±2.2% ( K i = 6.9 ±2.2 μ m ) β 2 AR in young rats and 66.1 ±1.2% ( K i = 8.3 ±1.1 nM) β 1 AR and 33.9 ±1.2% ( K i = 5.2 ± 0.6 μ m ) β 2 AR in old rats. The β 1 AR/β 2 AR ratio was significantly ( P <0.01) reduced in old rats with respect to the younger ones 4 By combining electrophysiological and binding measurements, we calculated β 1 AR and β 2 AR densities as number of receptors per μm 2 of cell surface. In old rats, β 1 AR density was significantly decreased compared to young rats (8.4±2.0 vs 15.4±3.7 receptors μm −2 , P <0.05), while β 2 AR density remained unchanged at both 3 and 22 months (3.8 ±0.7 and 4.2 ±1.1 receptors μm −2 , respectively) 5 Our results demonstrate that both β 1 AR and β 2 AR are functionally present in rat ventricular myocytes of young and old rats. The decreased responsiveness to βAR stimulation during aging appears to be associated with a selective reduction in the density of β 1 AR.