Angelo Forlani

Blockade of adenosine A2A receptors by SCH 58261 results in neuroprotective effects in cerebral ischaemia in rats.

BLOCKADE of adenosine receptors can reduce cerebral infarct size in the model of global ischaemia. Using the potent and selective A2A adenosine receptor antagonist, SCH 58261, we assessed whether A2A receptors are involved in the neuronal damage following focal cerebral ischaemia as induced by occluding the left middle cerebral artery. SCH 58261 (0.01 mg/kg either i.p. or i.v.) administered to normotensive rats 10min after ischaemia markedly reduced cortical infarct volume as measured 24 h later (30% vs controls, p < 0.05). Similar effects were observed when SCH 58261 (0.01 mg/kg, i.p.) was administered to hypertensive rats (28% infarct volume reduction vs controls, p < 0.05). Neuroprotective properties of SCH 58261 administered after ischaemia indicate that blockade of A2A adenosine receptors is a potentially useful biological target for the reduction of brain injury.

Characterization of the Potent and Highly Selective A2A Receptor Antagonists Preladenant and SCH 412348 [7-[2-[4-2,4-Difluorophenyl]-1-piperazinyl]ethyl]-2-(2-furanyl)-7H-pyrazolo[4,3-e][1,2,4]triazolo[1,5-c]pyrimidin-5-amine] in Rodent Models of Movement Disorders and Depression

The adenosine A2A receptor has been implicated in the underlying biology of various neurological and psychiatric disorders, including Parkinsons disease (PD) and depression. Preladenant and SCH 412348 [7-[2-[4-2,4-difluorophenyl]-1-piperazinyl]ethyl]-2-(2-furanyl)-7H-pyrazolo[4,3-e][1,2,4]triazolo[1,5-c]pyrimidin-5-amine] are potent competitive antagonists of the human A2A receptor (Ki = 1.1 and 0.6 nM, respectively) and have >1000-fold selectivity over all other adenosine receptors, making these compounds the most selective A2A receptor antagonists reported to date. Both compounds attenuate hypolocomotion induced by the A2A receptor agonist CGS-21680 [2-[p-(2-carboxyethyl)phenethylamino]-5′-N-ethylcarboxamidoadenosine], suggesting that they inhibit A2A receptor activity in vivo. Their high degree of selectivity and robust in vivo activity make preladenant and SCH 412348 useful tools to investigate the role of the A2A receptor system in animal models of PD and depression. Oral administration of preladenant and SCH 412348 (0.1–1 mg/kg) to rats potentiated 3,4-dihydroxy-l-phenylalanine (l-Dopa)-induced contralateral rotations after 6-hydroxydopamine lesions in the medial forebrain bundle and potently attenuated the cataleptic effects of haloperidol. Preladenant (1 mg/kg) inhibited l-Dopa-induced behavioral sensitization after repeated daily administration, which suggests a reduced risk of the development of dyskinesias. Finally, preladenant and SCH 412348 exhibited antidepressant-like profiles in models of behavioral despair, namely the mouse tail suspension test and the mouse and rat forced swim test. These studies demonstrate that preladenant and SCH 412348 are potent and selective A2A receptor antagonists and provide further evidence of the potential therapeutic benefits of A2A receptor inhibition in PD (with reduced risk of dyskinesias) and depression (one of the primary nonmotor symptoms of PD).

Lack of the nociceptin receptor does not affect acute or chronic nociception in mice

The peptide nociceptin/orphanin FQ (N/OFQ) and its receptor ORL-1, also designated opioid receptor 4 (OP(4)) are involved in the modulation of nociception. Using OP(4)-knockout mice, we have studied their response following opioid receptor stimulation and under neuropathic conditions.In vas deferens from wild-type and OP(4)-knockout mice, DAMGO (mu/OP(3) agonist), deltorphine II (delta/OP(1) agonist) and (-)-U-50488 (kappa/OP(2) agonist) induced similar concentration-dependent inhibition of electrically-evoked contractions. Naloxone and naltrindole (delta/OP(1) antagonists) shifted the curves of DAMGO (pA(2)=8.6) and deltorphine II (pA(2)=10.2) to the right, in each group. In the hot-plate assay, N/OFQ (10 nmol per mouse, i.t.) increased baseline latencies two-fold in wild-type mice while morphine (10mg/kg, s.c.), deltorphine II (10 nmol per mouse, i.c.v.) and dynorphin A (20 nmol per mouse, i.c.v.) increased hot-plate latencies by about four- to five-fold with no difference observed between wild-type and knockout mice. Furthermore, no change was evident in the development of the neuropathic condition due to chronic constriction injury (CCI) of the sciatic nerve, after both thermal and mechanical stimulation. Altogether these results suggest that the presence of OP(4) receptor is not crucial for (1) the development of either acute or neuropathic nociceptive responses, and for (2) the regulation of full receptor-mediated responses to opioid agonists, even though compensatory mechanisms could not be excluded.

Interaction of selected vasodilating β-blockers with adrenergic receptors in human cardiovascular tissues

β- And α1-adrenoceptor antagonist properties of bufuralol, carvedilol, celiprolol, dilevalol, labetalol, and pindolol were investigated in human myocardium and mammary artery using binding techniques and functional studies. In myocardial membranes, β-adrenoceptor antagonists showed monophasic competition isotherms for [l25I]pindolol binding with high affinity (Ki from 1–100 nM), except for celiprolol which displayed a bi-phasic competition isotherm (pKi = 6.4 ± 0.06 for pr and 4.8 ± 0.07 for β2-adrenoceptors). Drug interactions with α1-adrenoceptors were evaluated in human mammary artery by [3H]prazosin binding and by measuring contractile responses to norepinephrine (NE). Labetalol and carvedilol showed a moderate affinity for a,-adreno-ceptors (pKi = 6.2 ± 0.01 and 6.1 ± 0.06, respectively), and inhibited NE-induced contractions (pA2 = 6.93 ± 0.23 and 8.64 ± 0.24, respectively). Dilevalol, bufuralol, and pindolol displayed weak effect both in binding (Ki in micromolar range) and functional experiments (pA2 = 5.98, 5.54, and 6.23, respectively). Celiprolol did not show antagonist properties up to 100 μ.M in functional studies, but displayed a slight affinity for a,-adrenoceptors in binding studies. The data indicate that the vasodilating activity of these p-adrenoceptor antagonists is caused in some instances by an α1adrenoceptor antagonism (labetalol, carvedilol), whereas for the others alternative mechanisms should be considered.

Role of 5-HT2 receptors in serotonin-induced contraction in the human mammary artery

We studied the effects of serotonin (5-HT) on isolated human mammary arteries obtained from patients undergoing coronary by-pass grafting. 5-HT induced a concentration-dependent contractile response in the mammary artery, with an EC50 value of 0.34 microM. The 5-HT2 antagonist, ketanserin, reversed the contractions evoked by 5-HT in a competitive manner at a low concentration (10(-8) M), whereas non-competitive antagonism was apparent at higher concentrations (5 X 10(-8)-5 X 10(-7) M). To investigate whether the alpha 1-blocking component of ketanserin plays a role in the response observed in this vessel, we evaluated the effect of ketanserin on contractions induced by (-)-norepinephrine. Ketanserin, in concentrations up to 10(-7) M, did not influence the norepinephrine-induced contractions. Moreover, a threshold concentration of 5-HT (10(-7) M) amplified the contractile effect induced by norepinephrine (5 X 10(-8) M), and this response was inhibited by ketanserin (10(-7) M). The selective 5-HT3 antagonist, GR 38032F, did not affect the 5-HT-induced contractions. These findings indicate that the human mammary artery is a vascular tissue sensitive to 5-HT. The 5-HT2 receptor subtype appears to mediate the response.

Antihypertensive effect of spirapril and felodipine during repeated administration to spontaneously hypertensive rats

The antihypertensive activity of spirapril given alone or in combination with felodipine was investigated in spontaneously hypertensive rats (SHR) during a 3-week treatment regimen and for one week after drug withdrawal. Systolic blood pressure and heart rate were recorded once a week just before dosing and at varying time intervals up to 6 hr thereafter. Recordings were continued for one week after drug withdrawal. Spirapril alone at 1 and 5 mg/kg p.o. was found to produce dose-related antihypertensive effects throughout the treatment period. Felodipine alone at 5 mg/kg p.o. reduced blood pressure slightly more than did the low dose of spirapril. The combination of spirapril and felodipine induced a marked antihypertensive response which was greater than that observed in rats treated with either drug alone. One week after treatment withdrawal, blood pressure was at initial levels with no evidence of rebound phenomena. No significant heart rate changes were observed in the treated groups, as compared with the controls, except for an increase on the 1st day of treatment in rats given felodipine. These findings indicate that the combination of an angiotensin converting enzyme (ACE) inhibitor with a calcium antagonist leads to an effective control of hypertension over a prolonged period of treatment. Since the combination allows effectiveness with lower doses of ACE inhibitor, it is expected that the antihypertensive efficacy might be associated with a lower liability to untoward effects.

5-Hydroxytryptamine induces contraction in isolated human mammary artery: Effect of ketanserin

Summary5-hydroxytryptamine (5HT) treatment produced dose-related contractions in the human internal mammary artery with an EC50 value of 3.4×10-7M. The 5HT2 receptor antagonist ketanserin reversed the contractions evoked by 5HT in a competitive manner at a low concentration (10-6 M), whereas a noncompetitive antagonism was apparent at higher concentrations (5× 10-8 M to 5×10-7 M). The alpha1-blocking component of ketanserin was evaluated by studying the effect of ketanserin upon the contractile response evoked by norepinephrine. Up to 10-7 M, ketanserin did not influence norepinephrine-induced contractions. These findings indicate that the mammary artery is a vascular tissue sensitive to contractions induced by 5HT and that the drug ketanserin antagonizes this contractile response throuth the 5HT2 receptor subtype.