Adenosine A₁‐receptor stimulated increases in intracellular calcium in the smooth muscle cell line, DDT₁MF‐2

Abstract

1 The effect of a range of adenosine receptor agonists on intracellular free calcium concentration ([Ca2+]j) has been studied in the hamster vas deferens smooth muscle cell line DDTMF-2.2 Adenosine receptor agonists elicited a rapid and maintained increase in [Ca2+], in fura-2 loaded DDTIMF-2 cells. The initial rise could be maintained in the absence of extracellular calcium, whereas the maintained or plateau phase was dependent upon the presence of extracellular calcium and appeared to be associated with calcium influx. The rank order of agonist potencies was N6-cyclopentyladenosine > 5'-N-ethylcarboxamidoadenosine > 2-chloroadenosine > adenosine. 3 The response to 2-chloroadenosine was antagonized by the antagonists 8-cyclopentyl-1,3-dipropylxanthine (DPCPX, KD 0.14nM) and 8-phenyltheophylline (KD 112 nM).4 Pretreatment with the 5-lipoxygenase inhibitor AA861 (20 jLM) produced only a small (14 ± 2%) inhibition of the [Ca2+]i response elicted by N6-cyclopentyladenosine (300 nM), in nominally Ca2'-free buffer containing 0.1 mM EGTA. The cyclo-oxygenase inhibitor, indomethacin (2 pM) was without effect.5 The Ca2"-influx associated with the plateau phase required the continued presence of agonist on the receptor. The antagonist DPCPX (100 nM) attenuated the rise in [Ca2+]1 observed when extracellular Ca was re-applied after the cells had been stimulated with N6-cyclopentyladenosine (CPA;300 nM) in experiments initiated in nominally Ca2+-free buffer. 6 Pretreatment with pertussis toxin (200 ng ml-' for 4 h) inhibited the CPA (100 nM) stimulated intracellular Ca2+ release and Ca2" influx but was without effect on the response to histamine (100 fiM).7 These data suggest that adenosine Al-receptor activation in DDT1MF-2 cells stimulates release of Ca2+ from intracellular stores and influx of extracellular Ca2+ through Ca2+ entry pathways in the plasma membrane which required the continued presence of agonist on the receptor.

Adenosine A1Receptor-Mediated Activation of Phospholipase C in Cultured Astrocytes Depends on the Level of Receptor Expression

“…Like others (El-Etr et al, 1992;Dickenson and Hill, 1993;Akbar et al, 1994;Freund et al, 1994), we found that PTX completely abolished A 1 receptor potentiation of PLC activity, confirming an active role of G i /o -proteins in this process. There is no evidence that ␣ subunits of G i /o -proteins have any effects on PLC (Neer, 1995), but it is widely accepted that ␤␥ subunits are the signal transducers for the PTX-sensitive PLC stimulation mediated by G i /o -proteins (Zhu and Birnbaumer, 1996).…”

“…Very often the coupling of adenosine A 1 receptors to PLC␤ is synergistic, with the stimulation evoked by other receptors like ␣ 1 adrenergic receptors (El-Etr et al, 1989;Biber et al, 1996), histamine H 1 receptors (Dickenson and Hill, 1993), muscarinic receptors (Biden and Browne, 1993), or thyrotropin receptors (Okajima et al, 1995). There are, however, also reports suggesting direct activation of PLC by adenosine A 1 receptors (Gerwins and Fredholm, 1992b;Freund et al, 1994).…”

Coupling of an endogenous 5‐HT1B‐like receptor to increases in intracellular calcium through a pertussis toxin‐sensitive mechanism in CHO‐K1 cells

“…However, the pKb for methiothepin obtained in this study is about 1 order of magnitude higher than its affinity for rat and mouse 5-HTIB receptors (Boess & Martin, 1994 and OC2-adrenoceptors expressed at high concentrations in transfected CHO cells are able to inhibit adenylyl cyclase (the normal effector system for these receptors) and activate phospholipase C (Felder et al, 1991;Dell'Acqua et al, 1993;Iredale et al, 1994). In DDTIMF-2 cells, activation of the endogenously expressed adenosine Al receptor results in both the inhibition of adenylyl cyclase and the activation of phospholipase C through PTX-sensitive pathways (Ramkumar et al, 1990;White et al, 1992;Dickenson & Hill, 1993). Furthermore, Ramkumar et al (1990) have shown that the endogenous adenosine Al receptor in DDT1MF-2 cells is expressed at high levels (0.8-0.9 pmol mg-' of protein).…”

A1 adenosine receptor-induced phosphorylation and modulation of transglutaminase 2 activity in H9c2 cells: A role in cell survival

“…Furthermore of the five PKC isozymes expressed in H9c2 cells (PKC-, PKC-β1, PKC-, PKC-, and PKC-ζ) adenosine stimulates the nuclear translation of PKC- [50]. Since DAG activates PKC- and PKC-, it is conceivable that the Gi/o-protein coupled A1 adenosine receptor promotes PKC-/ activation via G-protein β subunit-mediated stimulation of phospholipase C [42,43,46]. It would be of interest in future studies to determine the PKC isozyme(s) involved in A1 adenosine receptorinduced TG2 activation.…”