N-[9-chloro-2-(2-furanyl)[1,2,4]-triazolo[1,5-c]quinazolin-5-benzeneacetamide (MRS-1220) and N6-3-iodobenzyl-5-N-methylcarbamoyladenosine (IB-MECA) were supplied by Tocris, U.K. (8-PT, 3 M), failed to antagonize these contractions, suggesting that A1/A2adenosine receptors were not involved. Unlike adenosine, R-PIA (3 M) produced contractions in non-incubated (0.230.04 g) or Krebs-incubated (0.150.04 g) tracheae, as well as after passive and active sensitization. None of these responses were blocked by 8-PT. The A3receptor agonist, IB-MECA, in the presence of 8-PT produced small contractions in passively sensitized tracheae (10 M, 0.020.003 g) and, in larger doses (100 Mand 1 mM), contracted actively sensitized tracheae. In actively sensitized trachea, the A3receptor antagonist, MRS-1220 (100 nM), significantly (P<0.05) attenuated adenosine contractions in the presence of 8-PT from 0.230.07 g to 0.070.03 g. These results show that passive, like active sensitization, reveals bronchoconstrictions to adenosine of isolated tracheae. The insensitivity GW6471 to 8-PT blockade, the antagonism by MRS-1220, and the fact that this A3receptor agonist, IB-MECA, mimics GW6471 this response, suggest involvement of A3receptors. R-PIA, however, has a different profile of adenosine receptor activity. Keywords:Adenosine, guinea-pig tracheal spirals, ovalbumen, passive sensitization, R-PIA, bronchoconstriction == Introduction == Inhaled adenosine (or 5-AMP) has little effect in normal subjects but causes bronchoconstriction in asthmatic and atopic non-asthmatics (Cushleyet al., 1983). Similarly, inhalation of adenosine by sensitized guinea-pigs causes bronchoconstriction but non-sensitized animals show no response (Thorne & Broadley, 1994;Spruntulis & Broadley, 2001). Thein vitroresponse of the airways to adenosine has been studied in isolated tissues from both sensitized and non-sensitized guinea-pigs. Adenosine usually causes a relaxation of non-sensitized guinea-pig tracheas (Brown & Collis, 1982;Darmani & Broadley, 1986). However,Advenieret al. (1982),Karlssonet al. (1982)andFarmeret al. (1988)have reported that adenosine can cause a small contraction of non-sensitized guinea-pig tracheas. In contrast,Pauwels & van der straeten (1987),Thorne & Broadley (1992),Lewiset al. (1994)andThorneet al. (1996)have shown that adenosine causes a more substantial contraction of the ovalbumen sensitized guinea-pig isolated airways. The mechanism responsible for this phenomenon is not comprehended. The bronchoconstriction in asthmatics can be inhibited by theophylline, an adenosine GW6471 receptor antagonist (Cushleyet al., 1983).Bjorcket al. (1992)have shown enhanced bronchoconstrictor responses to adenosine of isolated bronchi from asthmatics, which are mediated via adenosine A1receptors. It has therefore been suggested byAliet al. (1994)that there may be increased expression of A1receptors in the asthmatic airways. However,Walkeret al. (1997)reported that A3receptor expression was increased in asthmatic lung eosinophils. There are conflicting reports on which adenosine receptor mediates the adenosine-induced bronchoconstriction.El-Hashimet al. (1996)andGhaiet al. (1987)claim an A1receptor involvement, whereasThorne & Broadley (1992)andKehoe & Broadley (1996)suggest that A3receptors mediate the response in sensitized guinea-pigs. Recently,Hannonet al. (2002)have suggested that a novel atypical receptor, distinct from the four known categorized adenosine receptors, mediates the bronchoconstriction in sensitized rats. Sensitization of the animal to ovalbumen also results in a bronchoconstriction of guinea-pig airways to ovalbumenin vivo(Howellet al., 1993;Thorne & Broadley, 1994;Spruntulis & Broadley, 2001) andin vitro(Thorne & Broadley, 1992;Lewiset al., 1994;Broadley, 1995;Chabot-Fletcheret al., 1995). Active sensitization of guinea-pigs has long been used as a technique for the study of airway responsiveness and inflammationin vivoafter challenge of the animal with specific allergens (Busseet al., 1993;Pretolaniet al., 1994;Danahay & Broadley, 1997). In active sensitization, animals are pretreated with a sensitizing medium to raise antibodies to the particular antigen, in our experiments ovalbumen, together with the GW6471 adjuvant, aluminium hydroxide. In the procedure described for guinea-pigs byAndersson (1980), the immunoglobulins that were raised were of the E and G types. Exposure of sensitized airways to the specific antigenin vitroresults in an anaphylactic response, the release of mast cell mediators and a contractile response (Metcalfeet al., 1997). Passive sensitization by incubation of isolated human lung tissue with non-sensitized or sensitized serum is an alternative sensitization process (Tunon de laraet al., 1995), which has been used in a number of GW6471 studies to investigate the mediators and mechanisms involved in allergic reactions (Rabe, 1998;Schmidtet al., 2000). In passive sensitization, the isolated tissue from a Rabbit polyclonal to LACE1 non-sensitized subject is usually incubated with serum made up of elevated levels.