Influence of extracellular calcium and nifedipine on alpha 1- and alpha 2-adrenoceptor-mediated contractile responses in isolated rat and cat cerebral and mesenteric arteries

The influence of extracellular Ca2+ and nifedipine on contractile responses to 10 microM noradrenaline (NA) was investigated in isolated rat and cat middle cerebral (RCA, CCA) and mesenteric (RMA, CMA) arteries. In the CCA (containing predominantly alpha 2-adrenoceptors), the NA-induced contractions developed considerably more slowly than in the RCA, RMA (containing mainly alpha 1-adrenoceptors) and CMA (sensitive to both alpha 1- and alpha 2-adrenoceptor selective antagonists). The tonic component of the NA-induced contraction in the four types of artery was substantially suppressed after only short periods in Ca2+-free solution. In each type of artery, excluding the CCA, the contractile response to 124 mM K+ was more sensitive to Ca2+ deprivation than that to NA. This suggests that NA, besides mobilizing extracellular Ca2+, can also release Ca2+ from an intracellular pool in the RCA, RMA and CMA, but not in the CCA. Thus, alpha 1-adrenoceptor-mediated contractions in the RCA and RMA seem to depend on both Ca2+ influx and intracellular Ca2+ release, whereas alpha 2-adrenoceptor-mediated contractile responses in the CCA appear to rely almost entirely on Ca2+ influx. Both the maximum response and the tonic component of the NA-induced contraction were significantly more sensitive to nifedipine in the CCA than in the RCA. In comparison with the NA-induced contractions in these arteries, those in the RMA and CMA were relatively resistant to nifedipine. In the CCA exposed to NA in Ca2+-free medium, nifedipine almost abolished the contraction induced by re-addition of Ca2+, whereas in the other types of artery, Ca2+ re-application evoked a significant contraction also in the presence of the drug. The differential effects of nifedipine presumably reflect differences between the arteries, not only in the relative contribution of Ca2+ influx and intracellular Ca2+ release to the contractile activation, but also in the nifedipine sensitivity of the Ca2+ entry pathways utilized by NA. It is concluded that the mechanisms through which NA induces contraction seem to be related both to the subtype of alpha-adrenoceptor stimulated by NA and to the type of vessel studied.     

Effects of nimodipine,Bay K 8644 and pinacidil on α1- and α2-adrenoceptor-mediated vasoconstriction in human hand veins

The effects of nimodipine, Bay K 8644 and pinacidil, three drugs interfering with transmembrane Ca²⁺ fluxes in different ways, were investigated in isolated human hand veins. Their ability to influence the concentration-response relationship for noradrenaline (NA) was assessed in the absence and presence of prazosin or rauwolscine. The contractile response to NA was almost abolished in Ca²⁺-free medium. Nimodipine and pinacidil depressed the NA concentration-response curve both in the absence and presence of α-adrenoceptor blockers. The NA response was only partially inhibited by nimodipine, indicating that NA may activate nimodipine-insensitive influx pathways, presumably receptor-operated calcium channels. Pinacidil inhibited the contractile response to 124 mM K⁺ and reduced the NA-induced contraction in the presence of nimodipine, suggesting that pinacidil has actions other than the opening of potassium channels and subsequent membrane hyperpolarization. Bay K 8644 increased the NA potency fourfold in the presence of rauwolscine, whereas it had no effect on the NA response in the presence of prazosin and in the absence of α-adrenoceptor blockade. Such an action of Bay K 8644 can be reconciled with α-adrenoceptor activation causing membrane depolarization and opening of potential-operated calcium channels. It may be concluded that both α₁- and α₂-adrenoceptor-mediated contractions in human hand veins are highly dependent on Ca²⁺ influx, although the mechanisms utilized to bring about this influx partly differ between the two receptor subtypes.     

Reversal of α1-receptor mediated relaxation in intestinal smooth muscle

The α₁-receptor agonist phenylephrine relaxed longitudinal rabbit jejunal muscle contracted in vitro by low concentrations of barium ions (1 mM). When the Ba²⁺ concentration was increased to 10–15 mM the response to phenylephrine was a contraction, and at Ba²⁺ concentrations in between the high and low range this response was biphasic—a relaxation followed by a contractile phase. The α₂-receptor agonist clonidine did not affect the tone of the Ba²⁺ contracted preparation. When the muscle preparation was contracted by Sr²⁺ (1–20 mM) in the presence of Ca²⁺ (2.5 mM), phenylephrine relaxed it, and no contractile response to phenylephrine was observed. In the absence of extracellular Ca²⁺, 5 mM Ba²⁺ caused a contraction. Under these conditions phenylephrine had no effect on the tissue tone. When Ca²⁺ was added in a low concentration (0.2-2 mM), phenylephrine elicited a gradually increasing contractile response. At 5 mM Ca²⁺ the contractile response was replaced by the normal relaxation. The contractile response to phenylephrine in the presence of 5 mM Ba²⁺ and 2.5 mM Ca²⁺ was partially blocked by low concentrations of verapamil. In higher concentrations verapamil abolished the tissue tonus completely. The contractile response to phenylephrine in the presence of 5 mM Ba²⁺ and 2.5 mM Ca²⁺ could be reverted to the normal relaxation by the addition of 20 mM Mg²⁺. Increasing the K⁺ concentration from the normal 5.9 to 62.9 mM blocked the phenylephrine-induced relaxation. No contractile response to phenylephrine occurred. It is concluded that Ba²⁺ could reverse the response of α₁ receptor stimulation in rabbit jejunum from a relaxation to a contraction and that this contractile response was dependent on the presence of Ca²⁺.     

Cyclic AMP-mediated inhibition of noradrenaline-induced contraction and Ca2+ influx in guinea-pig vas deferens

The relaxation effects of forskolin and methylxanthines on noradrenaline (NA)-induced contractions were investigated by measuring isotonic contraction and intracellular calcium concentration ([Ca2+]i) in the epididymal side of guinea-pig vas deferens. NA (100 microM) and high K+ (55 mM) induced a biphasic contraction; fast, transient (phasic) and slow, sustained (tonic) phases. Both phases in either NA or high K+ stimulation were abolished in Ca2+-free solution. Pretreatment with 10 microM nifedipine, an L-type Ca2+ channel blocker, reduced both phasic and tonic contractions induced by high K+. In the case of NA-induced contraction, however, nifedipine reduced the phasic contraction but not the tonic contraction. The nifedipine-insensitive tonic contraction was relaxed by the application of polyvalent cations (Mn2+, Co2+, Cd2+ and La3+). These findings indicate that NA-induced biphasic contraction is mainly due to nifedipine-insensitive Ca2+ influx, especially in the tonic phase. Cyclic AMP-increasing agents such as forskolin (0.5-10 microM), IBMX (5-500 microM) and caffeine (1-20 mM) relaxed the NA-induced contraction extensively in a concentration-dependent manner. However, these agents only partially relaxed the high K+-induced contraction. Forskolin (10 microM) and IBMX (100 microM) reduced the [Ca2+]i response to NA, but had no effect on the [Ca2+]i response to high K+. These results suggest that an increase in intracellular cAMP may relax the NA-induced contraction by attenuating a nifedipine-insensitive Ca2+ influx and by a mechanism independent of a reduction in [Ca2+]i.     

Postjunctional α1- and α2-adrenoceptors mediating contraction in isolated human groin arteries and veins

By means of selective agonists and antagonists for α₁- and α₂-receptors, the α-receptor subtypes in human groin arteries and veins were characterized and compared. In the arteries the α₁-receptor blocker prazosin caused a concentration-dependent parallel displacement of the noradrenaline (NA) concentration-response (cr) curve without reduction of maximum (pA₂=9.86); the selective α₂-receptor antagonist rauwolscine in the concentration 10^-8 M caused a right-ward shift of the NA cr-curve without reduction of E_max, but 10^-7 M and 10^-6 M caused little or no further shift. In the veins, the two antagonists had the opposite effects. Rauwolscine caused a concentration-dependent right-ward shift of the NA cr-curve without depression of maximum (pA₂=9.03); prazosin 10^-9 M significantly displaced the NA cr-curve, whereas 10^-8 M and 10^-7 M caused little or no further shift. The responses to the α₂-receptor agonist clonidine in the arteries were too small to allow calculations of pEC50 values; in the veins contractions were elicited in all vessel segments investigated (pEC₅₀=6.24). Phenylephrine, selective for α₁-receptors, was significantly more potent in arteries than in veins. NA was significantly more potent in veins than in arteries. It is concluded that in human groin vessels, there is a functional predominance of arreceptors in the arteries and of a2-receptors in the veins.     

Characterization of two different calcium entry pathways in small mesenteric arteries from rat

The effects of Ca2+ removal, nifedipine, and La3+ on contractions induced by 124 mM K+ and 10 microM noradrenaline (NA) were investigated in small mesenteric arteries from rat. Ring segments of the arteries were suspended between two steel wires in a 2.5 ml muscle bath, and the mechanical activity recorded "isometrically". The tonic components of the contractile responses to both K+ and NA were critically dependent on the presence of Ca2+ in the bath solution. Nifedipine effectively relaxed K+-contracted arteries, whereas those activated by NA were considerably less affected by the drug. Application of NA to arteries depolarized by K+ in the presence of nifedipine induced a sustained tonic contraction, which was only approximately 20% smaller than that elicited by NA in "standard" Krebs solution, implicating pharmacomechanical coupling. Unlike nifedipine, La3+ inhibited K+- and NA-induced contractions to approximately the same extent. Re-application of Ca2+ to "Ca2+-depleted" preparations exposed to K+ and/or NA induced concentration-dependent contractions. The experimental results suggested that the effects of K+ and NA on the membrane permeability to Ca2+ were additive. The Ca2+-induced contractions were more inhibited by nifedipine in K+-depolarized than in NA-exposed arteries. It is concluded that K+ and NA utilize partly different Ca2+ entry pathways to increase the myoplasmic Ca2+ concentration in rat mesenteric arteries. Whereas K+ seems to promote the influx Ca2+ by activation of CA2+ channels sensitive to the membrane potential, the nature of the receptor-operated Ca2+ entry pathway remains to be established.     

Actions of angiotensin II and noradrenaline on smooth muscle cells of the canine mesenteric artery

The actions of angiotensin II (AngII) and noradrenaline (NA) on smooth muscle cells of the canine mesenteric artery were studied by measurement of isometric contractions recorded from muscle strips and the intracellular Ca²⁺ concentration monitored with quin2-fluorescence from dispersed suspensions of single cells. The Ca²⁺ transients provoked by the two agonists were monophasic in shape, i.e., after application of each agonist, [Ca²⁺]_i rose immediately within 1 s and decreased to near-basal level within 5 min. The contraction induced by NA was maintained for several minutes whilst that induced by AngII was short-lasting. When NA was repetitively applied to the strip in Ca²⁺-containing solution, the same amplitude of contractions was always obtained. In contrast, after initial exposure to AngII, subsequently-applied AngII generated small contractions. In Ca²⁺-free solution, either agonist could induce the large contraction. After initial exposure to NA or AngII in Ca²⁺-free solution, subsequently-induced contractions by either agonist were reduced. The response induced by AngII was blocked by [Sar¹, Ile⁸]-AngII and that of NA was blocked by phentolamine. Pertussis toxin inhibited contractions induced by both agonists but not those induced by caffeine and high K⁺. An activator of protein kinase C, 12-O-tetradecanoylphorbol 13-acetate (TPA), produced a slowly-developing contraction without any change in [Ca²⁺]_i, and this agent inhibited the contractions and Ca²⁺ transients induced by both agonists. These results indicate that NA and AngII each act on a specific receptor and release Ca²⁺ from common intracellular storage sites through production of inositol 1,4,5-trisphosphate (InsP₃). However, the actions of protein kinase C or the Ca²⁺-releasing mechanisms induced by InsP₃ do not seem to be causally related to the generation of homologous desensitization induced by AngII.     

Inhibition of PKC-dependent extracellular Ca2+ entry contributes to the depression of contractile activity in long-term pressure-overloaded endothelium-denuded rat aortas

We examined the contractile responsiveness of rat thoracic aortas under pressure overload after long-term suprarenal abdominal aortic coarctation (lt-Srac). Endothelium-dependent angiotensin II (ANG II) type 2 receptor (AT₂R)-mediated depression of contractions to ANG II has been reported in short-term (1 week) pressure-overloaded rat aortas. Contractility was evaluated in the aortic rings of rats subjected to lt-Srac or sham surgery (Sham) for 8 weeks. ANG I and II levels and AT₂R protein expression in the aortas of lt-Srac and Sham rats were also evaluated. lt-Srac attenuated the contractions of ANG II and phenylephrine in the aortas in an endothelium-independent manner. However, lt-Srac did not influence the transient contractions induced in endothelium-denuded aortic rings by ANG II, phenylephrine, or caffeine in Ca²⁺-free medium or the subsequent tonic constrictions induced by the addition of Ca²⁺ in the absence of agonists. Thus, the contractions induced by Ca²⁺ release from intracellular stores and Ca²⁺ influx through stored-operated channels were not inhibited in the aortas of lt-Srac rats. Potassium-elicited contractions in endothelium-denuded aortic rings of lt-Srac rats remained unaltered compared with control tissues. Consequently, the contractile depression observed in aortic tissues of lt-Srac rats cannot be explained by direct inhibition of voltage-operated Ca²⁺ channels. Interestingly, 12-O-tetradecanoylphorbol-13-acetate-induced contractions in endothelium-denuded aortic rings of lt-Srac rats were depressed in the presence but not in the absence of extracellular Ca²⁺. Neither levels of angiotensins nor of AT₂R were modified in the aortas after lt-Srac. The results suggest that, in rat thoracic aortas, lt-Srac selectively inhibited protein kinase C-mediated activation of contraction that is dependent on extracellular Ca²⁺ entry.     

Kinetics of ATP- and noradrenaline-mediated sympathetic neuromuscular transmission in rat tail artery

Electrophysiological, electrochemical and mechanical recordings were employed to study the kinetics of the release and clearance of adenosine 5′-triphosphate (ATP) and noradrenaline (NA) as sympathetic co-transmitters and of the neurogenic and non-neurogenic contractions in rat isolated tail artery. The life-time of ATP and NA released by a single pulse or 10 pulses at 50 Hz was brief (< 100 ms, or < 3 s, respectively); the neurogenic contractile responses occurred largely after the transmitters had been removed from the extracellular space. The ATP-induced neurogenic contractile responses to a single pulse or 10 pulses at 50 Hz were similar in time-course to the responses to direct muscle stimulation at low voltage; both seemed to be caused by activation of nifedipine-sensitive voltage-gated L-type Ca²⁺ channels. The a,- and a₂-adrenoceptor-mediated components of the NA-induced neurogenic contractile response to 10 pulses at 50 Hz were more delayed and prolonged and determined by properties of the post-receptor mechanisms. The per pulse release of both ATP and NA faded rapidly during long high-frequency trains. So did the ATP level at the receptors and the ATP-induced neurogenic contraction. The NA levels and the contractile responses induced via a,- and ^-adrenoceptors were much better maintained during ongoing stimulation at 20 Hz but relaxed rapidly afterwards, suggesting that nerve activity suppressed, and ce'ssation of nerve activity reactivated NA clearance.     

Affinity of Noradrenaline and Dopamine for Neural α-Receptors Mediating Negative Feedback Control of Noradrenaline Secretion in Human Vasoconstrictor Nerves

Isolated superfused field stimulated biopsy specimens of human peripheral arteries and veins, preincubated with ³H-(-)-noradrenaline (NA) to label the neural stores of NA, were used to study the potency of dopamine (DA) and of NA as triggers of α-adrenoceptor mediated negative feedback control of sympathetic neurotransmitter secretion, evoked by stimulation with trains of 300 shocks at 1 Hz. In this preparation DA was found to be only slightly less potent than NA in depressing both the secretion of ³H-NA, and the contractile response, evoked by nerve stimulation. DA depressed the contraction evoked by exogenous NA as well, but to a very much smaller extent. On the other hand, DA was a very weak agonist on the α-receptors of the smooth muscle; nearly 1 000 times higher concentrations of DA were required to mimick contractions evoked by exogenous NA. The results show that the neural α-receptor function involved in control of NA secretion differs considerably from the α-receptors of e.g. smooth muscle, with respect to sensitivity to DA. It seems possible that the observed depressing effect of DA on NA secretion may be of pharmacological and clinical interest; it may at least in part explain the vasodilating effect of DA infusions in man.     

Pharmacological properties of prejunctional α-adrenoceptors in isolated feline middle cerebral arteries; comparison with the postjunctional α-adrenoceptors

In cat middle cerebral arteries (CMCA) preincubated with ³H-noradrenaline (NA), the outflow of tritium evoked by electrical stimulation was reduced to 32% by α-adrenoceptor (α-receptor) stimulation with oxymetazoline, and increased to 487% by α-receptor blockade with HEAT. The relative order of potency for α-receptor agonists on prejunctional receptors was: clonidine ≥ oxymetazoline > phenylephrine, and the antagonist rauwolscine was more potent than prazosin. This indicates that the prejunctional α-receptors are mainly of α₂-type. Rauwolscine was more potent than prazosin in inhibiting the contractions induced by NA, indicating a predominance of α₂-receptors postjunctionally. Apart from clonidine having higher intrinsic activity pre- than postjunctionally, all drugs examined (oxymetazoline, phenylephrine, rauwolscine, HEAT (BE2254), and prazosin) had similar concentration-effect curves on the pre- and postjunctional receptors. Furthermore, the ratios of EC₅₀-values pre- and postjunctionally of rauwolscine, oxymetazoline, and clonidine were all close to unity. These results indicate that pre- and postjunctional α₂-receptors in the CMCA have similar pharmacological characteristics and cannot be influenced separately by the presently used drugs.     

α2-Aclrenoceptor agonist-mediated inhibition of [3H]noradrenaline release from rat hippocampus is reduced by 4-aminopyridine,but that caused by an adenosine analogue or co-conotoxin is not

The inhibitory effect of an adenosine analogue, R-PIA, and an α₂-adrenoceptor agonist, UK 14,304, on [³H]NA efflux from field-stimulated rat hippocampal slices was examined. The effect of 0.1 μm UK 14,304 was mimicked by 30 nM w-conotoxin and by 10 μM cadmium chloride, inhibitors of N- and L-type Ca²⁺ channels. R-PIA (1 μM) had no effect per se, but caused a clear-cut inhibition after blockade of the pre-synaptic α₂-receptor by yohimbine. 4-Aminopyridine (4-AP) caused a dose-dependent increase in evoked transmitter release. At 30 μM 4-AP did not affect the actions of ω-conotoxin or cadmium chloride. The pre-synaptic effect of R-PIA was similarly unaffected by 30 μM 4-AP. The presynaptic effect of UK 14,304 was virtually abolished by 4-AP (30 μM). The effect of UK 14,304 (0.1 μM) could be partly restored by reducing the Ca²⁺ concentration during treatment with 4-AP (22% inhibition compared to 42% with normal Ca²⁺). The magnitude of increase in evoked [³H]NA efflux by yohimbine (1 μM) was decreased by 4-AP in a concentration-dependent manner from 142% increase in controls to 21 % at 100 μM 4-AP. The present results indicate that NA release is reduced by somewhat different mechanisms by pre-synaptic α₂- and adenosine Aj-receptors. Furthermore, the results indicate that pre-synaptic A_rreceptors on hippocampal NA neurons do not primarily regulate 4-AP-dependent potassium channels, but they might act directly on a Ca²⁺ conductance.     

Effects of prostanoids on isolated feline cerebral arteries

The roles of extra-and intracellular calcium for the contractile effects of PGF_2α in the feline basilar artery (BA) were investigated. Comparisons were made with contractions induced by K⁺ and noradrenaline (NA). Addition of nifedipine to PGF_2α-or K⁺ (124 mM)-contracted arteries resulted in an incomplete relaxation, whereas NA-contracted vessels were completely relaxed. Incubation of the preparations in a calcium-free medium containing 10^-5 M EGTA for 5–10 min almost abolished contractions induced by K⁺ and NA. In contrast, 63 % of the response to PGF_2α remained after pretreatment of the arteries in a calcium-free solution for 40 min; PGF_2α produced a biphasic contraction in 17 out of 20 preparations consisting of a rapidly developing initial phase followed by a second increase in tension after 1–6 min. The second phase was absent if the EGTA-concentration was increased to 10^-4 M, or if the arteries were pre-treated with nifedipine. After incubation of the arteries in a calcium-free medium for 40–120 min and K⁺-depolarization, re-addition of calcium elicited contractions at lower concentrations in the presence of PGF_2α than in controls. The results suggest that PGF_2α-induced contractions in the feline BA are considerably less dependent on extracellular calcium than contractions evoked by K⁺ or NA. PGF_2α appears to be able to release calcium from two cellular stores, and may also promote calcium influx through the cell membrane.     

Effects of α- and β-adrenoceptor stimulation on 45Ca2+ efflux and insulin secretion from perfused rat islets

Exposure to the β₂-adrenoceptor agonist terbutaline resulted in a transient stimulation of ⁴⁵Ca²⁺ efflux from ⁴⁵Ca²⁺ preloaded rat islets perfused in 2 mm Ca²⁺ and 8.3 mm glucose. Concomitantly, an increase in insulin secretion occurred. Under the same experimental conditions, the α-adrenoceptor agonist noradrenaline promptly inhibited insulin release without any apparent influence on ⁴⁵Ca²⁺ efflux. In contrast, in a medium containing 2 mm Ca²⁺ and a low glucose concentration (2.8 mm), terbutaline stimulated insulin secretion without any apparent effects on ⁴⁵Ca²⁺ efflux. Noradrenaline had no effect on insulin secretion or ⁴⁵Ca²⁺ efflux in this medium. When islets were perfused with 8.3 mm glucose in a Ca²⁺ deficient medium, with or without addition of the chelating agent EGTA, terbutaline induced a marginal stimulation of insulin secretion and a negligible stimulation of ⁴⁵Ca²⁺ efflux. On the contrary, noradrenaline stimulated to an immediate and notable ⁴⁵Ca²⁺ efflux in these Ca²⁺ deficient media. Noradrenaline also clearly inhibited insulin secretion, though less markedly and with a slower onset than in islets perfused in 2 mm Ca²⁺. When the islets were perfused in a Ca²⁺ deficient medium with 2.8 mm glucose, terbutaline had a slight insulin releasing effect, but stimulated ⁴⁵Ca²⁺ efflux potently. Noradrenaline had no influence on insulin secretion but a weak stimulatory effect on ⁴⁵Ca²⁺ efflux. The data suggest that the β₂-adrenoceptor agonist terbutaline has the ability to stimulate insulin secretion in perfused rat islets, requiring extracellular Ca²⁺ for the full expression of its effects. These effects may be exerted through a Ca²⁺-Ca²⁺ exchange over the cell membrane and/or through cAMP and intracellular Ca²⁺ perturbations. Moreover, terbutaline directly stimulates ⁴⁵Ca²⁺ efflux, an effect inhibited by glucose. Further, the α-adrenoceptor agonist noradrenaline can inhibit insulin secretion in the absence of extracellular Ca²⁺, but the full expression of its inhibitory action is dependent on extracellular Ca²⁺ and glucose. In addition, noradrenaline stimulates ⁴⁵Ca²⁺ efflux in a Ca²⁺ deficient medium, an effect which appears independent of the glucose concentration.     

Effects of α1‐acid Glycoprotein Fucosylation on its Ca2+ Mobilizing Capacity in Neutrophils

We recently showed that the acute‐phase protein α₁‐acid glycoprotein (AGP) induces rises in cytosolic calcium concentration, [Ca²⁺]_i, in neutrophils through sialic acid dependent interactions with the neutrophil receptors siglec‐5 and/or siglec‐14. Whereas both siglec‐5 and siglec‐14 have a relatively broad specificity for sialylated oligosaccharide structures, including both structures with terminal α2–3 or α2–6 linked sialic acid, there is a markedly reduced affinity to the fucosylated epitope sialyl Lewis x (SLe^x). Increased fucosylation, leading to increased expression of SLe^x on AGP is commonly associated with inflammatory conditions. In the present study, we investigated whether an increased SLe^x expression would affect the Ca²⁺‐mobilizing effect of AGP. AGP with elevated fucose content isolated from patients with untreated chronic joint inflammation showed a decreased [Ca²⁺]_i modulatory effect on neutrophils compared to normally fucosylated AGP. Furthermore a hyperfucosylated AGP form produced by in vitro fucosylation, that consequently had an elevated expression of SLe^x, could not elicit a [Ca²⁺]_i increase in neutrophils. The role of the carbohydrate portion of AGP in modulating neutrophil responses was further strengthened by showing that synthetic glycoconjugates carrying oligosaccharides with terminal α2–3 or α2–6 linked sialic acid were able to mimic the Ca²⁺‐mobilizing effect of AGP whereas a synthetic glycoconjugate carrying SLe^x was not. Based on these data, we conclude that increased fucosylation can alter the ability of AGP to induce neutrophil signalling and further supports an important role of the oligosaccharide chains of AGP in the modulation of leukocyte functions during an inflammatory process.     

Modulation of vascular contractile responses to α1-and α2-adrenergic and neuropeptide Y receptor stimulation in rats with ischaemic heart failure

In order to evaluate adaptational changes in vascular function in congestive heart failure (CHF), we studied the contractile responses of isolated arterial and venous blood vessels from rats suffering from CHF induced by coronary artery ligature, resulting in a myocardial infarction. The contractile responses of the basilar, femoral and renal arteries and of the iliac vein were examined in relation to adrenergic and neuropeptide Y (NPY) receptor function by the action of the α₁ agonist phenylephrine, the α₂ agonist clonidine and NPY. The contractile force was measured (in mN) and in% of K⁺-induced contraction as well as pD₂ to each agonist. When stimulated by a 60 mM K⁺-buffer solution, the femoral and renal arteries from CHF rats responded with a stronger contraction (E_max; 9.4 ± 0.6 and 9.8 ± 0.6mN) than the corresponding Sham vessels (E_max; 6.2 ± 0.7 and 5.6 ± 0.4 mN respectively, P < 0.001). On the contrary, the iliac vein of CHF responded less to K⁺ than the Sham iliac vein (E_mas 2.5 ± 0.2 and 3.7 ± 0.5 mN, P < 0.01). The CHF iliac vein responded with a weaker contraction when stimulated with phenylephrine (E_max 1.9 ± 0.4 mN) and showed a lower sensitivity (pD₂ 5.6 ± 0.1) than the corresponding sham vessel (E_max 5.7 ± 2.3mN and pD₂ 6.3 ± 0.5, P < 0.05). The CHF renal artery was less sensitive to clonidine (pD₂ 6.4 ± 0.6) than the Sham renal artery (pD₂ 7.2 ± 0.1, P < 0.05). The results indicate differences between CHF and Sham vessel segments according to both contractile capacity induced by K⁺-depolarization and to agonist induced contractile capacity and sensitivity. The differences are not of general nature but vary according to the vascular bed examined.     

The role of sarcoplasmic reticulum and sarcoplasmic reticulum Ca2+-ATPase in the smooth muscle tone of the cat gastric fundus

Circular smooth muscle strips isolated from cat gastric fundus were studied in order to understand whether the sarcoplasmic reticulum (SR) and SR Ca²⁺-ATPase could play a role in the regulation of the muscle tone. Cyclopiazonic acid (CPA), a specific inhibitor of SR Ca²⁺-ATPase, caused a significant and sustained increase in muscle tone, depending on the presence of extracellular Ca²⁺. Nifedipine and cinnarizin only partially suppressed the CPA-induced tonic contraction. Bay K 8644 antagonized the relaxant effect of nifedipine in CPA-contracted fundus. Nitric-oxide-releasing agents sodium nitroprusside and 3-morpholino-sydnonimine completely suppressed the CPA-induced tonic contraction. The blockers of Ca²⁺-activated K⁺ channels, tetraethylammonium, charybdotoxin and/or apamin, decreased the contractile effect of CPA. Vanadate increased the tone but did not change significantly the effect of CPA. CPA exerted its contractile effect even when Ca²⁺ influx was triggered through the Na⁺/Ca²⁺ exchanger and the other Ca²⁺ entry pathways were blocked. Thapsigargin, another specific SR Ca²⁺-ATPase inhibitor, also increased the muscle tone. The effect of thapsigargin was completely suppressed by sodium nitroprusside and 3-morpholino-sydnonimine and partially by nifedipine. In conclusion, under conditions when the SR Ca²⁺-ATPase is inhibited, the tissue develops a strong tonic contraction and a large part of this is mediated by Ca²⁺ influx presumably via nifedipine-sensitive Ca²⁺ channels. This study suggests the important role of SR Ca²⁺-ATPase in the modulation of the muscle tone and the function of SR as a “buffer barrier” to Ca²⁺ entry in the cat gastric fundus smooth muscle. Received: 10 August 1995/Received after revision: 9 November 1995/Accepted: 10 November 1995     

Effects of nimodipine, Bay K 8644 and pinacidil on alpha 1- and alpha 2-adrenoceptor-mediated vasoconstriction in human hand veins

The effects of nimodipine, Bay K 8644 and pinacidil, three drugs interfering with transmembrane Ca2+ fluxes in different ways, were investigated in isolated human hand veins. Their ability to influence the concentration-response relationship for noradrenaline (NA) was assessed in the absence and presence of prazosin or rauwolscine. The contractile response to NA was almost abolished in Ca2+ -free medium. Nimodipine and pinacidil depressed the NA concentration-response curve both in the absence and presence of alpha-adrenoceptor blockers. The NA response was only partially inhibited by nimodipine, indicating that NA may activate nimodipine-insensitive influx pathways, presumably receptor-operated calcium channels. Pinacidil inhibited the contractile response to 124 mM K+ and reduced the NA-induced contraction in the presence of nimodipine, suggesting that pinacidil has actions other than the opening of potassium channels and subsequent membrane hyperpolarization. Bay K 8644 increased the NA potency fourfold in the presence of rauwolscine, whereas it had no effect on the NA response in the presence of prazosin and in the absence of alpha-adrenoceptor blockade. Such an action of Bay K 8644 can be reconciled with alpha 1-adrenoceptor activation causing membrane depolarization and opening of potential-operated calcium channels. It may be concluded that both alpha 1- and alpha 2-adrenoceptor-mediated contractions in human hand veins are highly dependent on Ca2+ influx, although the mechanisms utilized to bring about this influx partly differ between the two receptor subtypes.     

Recognition of E-cadherin on epithelial cells by the mucosal T cell integrin αM290β7 (αEβ7)

The integrin α_M290β7 on the surface of a T cell hybridoma, MTC-1, mediated adhesion of these cells to the mouse epithelial cell line CMT93. This interaction was critically dependent on the presence of divalent cations; Mn²⁺ strongly promoted adhesion, Ca²⁺ was ineffective and Mg²⁺ gave intermediate results. Antibodies to molecules on the surface of CMT93 cells were tested for inhibition of adhesion. One monoclonal antibody (mAb) against E-cadherin, ECCD-2, was found to have significant inhibitory activity. Other mAb to E-cadherin and antibodies to other molecules had no effect. To show that inhibition by ECCD-2 was specific for adhesion mediated by α_M290β7, MTC-1 cells were induced to adhere to CMT93 via the LFA-1/ICAM-1 pathway. For this purpose, the epithelial cells were treated with interferon-γ and tumor necrosis factor-α to induce ICAM-1 expression and, in addition, α_M290β7 on MTC-1 cells was down-regulated by culturing the cells in the absence of transforming growth factor β. Under these circumstances adhesion of MTC-1 cells to CMT93 was inhibited by an antibody to LFA-1 but not by ECCD-2. Transfection of mouse L cells with cDNA for mouse E-cadherin enabled MTC-1 cells to adhere to them through the α_M290β7 integrin; this interaction was inhibited both by ECCD-2 and by blocking antibody against the integrin. These data strongly suggest that E-cadherin is a principal ligand for α_M290β7.