Two typical properties of Gi o protein
Two typical properties of Gi/o protein-coupled receptors, namely an increase in 35S-GTPγS binding (Strange, 2010) and an inhibition of exocytotic noradrenaline release (Schlicker and Göthert, 1998), were not shared by H4 receptor activation on cortical membranes (guinea-pig and mouse) and cortical slices (human, guinea-pig and mouse), respectively, although activation of H3 receptors, which are Gi/o protein-coupled as well (Alexander et al., 2013), elicited both responses (Feliszek et al., 2015). By contrast, not only H3, but also H4, receptor activation inhibited noradrenaline release on guinea-pig cardiac synaptosomes in the study by Chan et al. (2012). The aim of the present study was to extend their work in a threefold manner. First, another preparation of the guinea-pig atrium (i.e. isolated tissue pieces) was chosen to confirm the inhibitory effect of H4 receptor activation. Second, we were interested whether an H4 receptor-mediated inhibition of noradrenaline release is detectable in another three sympathetically innervated tissues or organs, i.e. the aorta, renal cortex and vas deferens. Third, we examined whether H4 receptor activation, although not affecting noradrenaline release in the guinea-pig cortex (Feliszek et al., 2015), may inhibit the release of another transmitter, i.e. acetylcholine, in guinea-pig hippocampal slices. The effects of the H4 receptor agonist 4-methylhistamine and the H3 receptor agonist R-α-methylhistamine were studied in parallel. The muscarinic receptor agonist oxotremorine was used as a positive control since cyp3a inhibitors release in the brain and noradrenaline release in sympathetically innervated organs or tissues is strongly inhibited via presynaptic muscarinic auto- (Starke et al., 1989) and heteroreceptors (Boehm and Kubista, 2002, Kurz et al., 2008), respectively.
Material and methods
Results Experiments were performed on hippocampal slices preincubated with 3H-choline and on four sympathetically innervated periperal organs or tissues preincubated with 3H-noradrenaline. Basal tritium efflux was expressed as t1 or t2/t1 (see Methods). The t1 values in controls are given in Table 1; they were not affected by thioperamide 0.1 μM (not shown). The t2/t1 value was close to 0.8 in control experiments and not affected by the drugs under study (not shown). The electrically evoked tritium overflow was expressed as S1 or S2/S1 (see Methods). Control values are given in Table 1 (S1) and in Fig. 1 and the legend to Fig. 2 (S2/S1). The effects of the test drugs were examined in two experimental series shown in Fig. 1, Fig. 2. In the first series, the effect of the histamine H3 receptor agonist R-α-methylhistamine and the H4 receptor agonist 4-methylhistamine on the electrically evoked tritium overflow (S2/S1) was studied on hippocampal slices and sympathetically innervated organs or tissues. The muscarinic receptor agonist oxotremorine served as a positive control. The three drugs were administered at concentrations that elicit the maximum effect at the respective receptors. R-α-Methylhistamine inhibited the electrically evoked tritium overflow in tissue pieces from the aorta, atrium, renal cortex and vas deferens without affecting that in hippocampal slices (Fig. 1). 4-Methylhistamine failed to alter the evoked overflow whereas oxotremorine strongly inhibited it in each of the five tissues (Fig. 1). The second series of experiments was dedicated to the H3 receptors on the sympathetic neurones innervating the aorta and renal cortex. For this purpose, the interaction of histamine with thioperamide was examined. Histamine inhibited the electrically evoked tritium overflow (S2/S1) in the aorta and renal cortex in a concentration-dependent manner (Fig. 2). The maximum inhibitory effect of histamine was close to 50% in either tissue and its potency (pEC50) was 5.6 (aorta) and 5.9 (renal cortex). The concentration–response curve of histamine was shifted to the right by thioperamide 0.1 μM (Fig. 2), yielding apparent pA2 values of 8.0 (aorta) and 8.1 (renal cortex). Thioperamide 0.1 μM did not affect the evoked tritium overflow (S1) by itself (results not shown).