Jürgen Harting

A pharmacological profile of the novel, peripherally-selective k-opioid receptor agonist, EMD 61753

1 The pharmacological properties of the novel diarylacetamide k‐opioid receptor agonist, EMD 61753, have been compared with those of ICI 197067 (a centrally‐acting k agonist) and ICI204448 (a peripherally‐selective k agonist). 2 EMD 61753 binds with high affinity (IC50 5.6 nm) and selectivity (k:μ:δ:σ binding ratio 1:536: 125: > 1,786) to k‐opioid receptors and is a full and potent (IC50 54.5 nm) agonist in an in vitro assay for k‐opioid receptors (rabbit vas deferens preparation). 3 Systemically‐applied [14C]‐EMD 61753 is found in high concentrations in the lungs, liver, adrenal glands and kidneys. Considerably less radioactivity is detected in the whole brain, and this radioactivity is concentrated in the region of the cerebral ventricles in the choroid plexuses. EMD 61753 penetrates only poorly into the CNS. 4 EMD 61753 was weakly effective in pharmacological tests of central activity. This compound reversed haloperidolol‐induced DOPA accumulation in the nucleus accumbens of the rat only at a dose of 30mg kg−1, s.c., (doses of 0.1, 1.0 and 10mg kg−1, s.c., and 1.0, 10 and 100 mg kg−1, p.o., were inactive). Hexobarbitone‐induced sleeping in mice was prolonged by EMD 61753 at threshold doses of 10 mg kg−1, s.c., and 100 mg kg−1, p.o., whereas the motor performance of rats in the rotarod test was impaired by EMD 61753 with an ID50 value of 453 mg kg−1, s.c. 5 EMD 61753 produced dose‐dependent, naloxone‐reversible antinociception in the mouse formalin test (1st phase ID50 1.9 mg kg−1, s.c., and 10.4 mg kg−1, p.o.; 2nd phase ID50 0.26 mg kg−1, s.c., and 3.5 mg kg−1 p.o.) and rodent abdominal constriction test (ID50 mouse 1.75 mg kg−1, s.c., and 8.4 mg kg−1, p.o.; ID50 rat 3.2 mg kg−1, s.c., and 250 mg kg−1, p.o.). EMD 61753 was inactive, or only weakly effective, in the rat pressure test under normalgesic conditions. After the induction of hyperalgesia with carrageenin, however, this compound elicited potent, dose‐dependent (ID50 0.08 mg kg−1, s.c., and 6.9 mg kg−1, p.o., after remedial application, and 0.2 mg kg−1, s.c., and 3.1 mg kg−1, p.o., after prophylactic application) and naloxone‐reversible antinociception. The antinociceptive action of systemically‐applied (50 mg kg−1, p.o.) EMD 61753 in the hyperalgesic pressure test was completely inhibited by injection of the k‐opioid antagonist norbinaltorphimine (100 μg) into the inflamed tissue, a result which indicates that this opioid effect is mediated peripherally. 6 Cutaneous plasma protein extravasation produced by antidromic electrical stimulation of the rat saphenous nerve was dose‐dependently inhibited by systemically‐applied EMD 61753 (ID50 values 3.7 mg kg−1, s.c., and 35.8 mg kg−1, p.o.), and this effect was completely antagonized by intraplantar application of norbinaltorphimine (50 μg). Extravasation elicited by the intraplantar application of substance P (10 μg) was not influenced by the administration of EMD 61753. 7 EMD 61753 produced dose‐dependent diuresis in non‐hydrated rats at doses of and above 10 mg kg−1, s.c., and 10 mg kg−1, p.o., and in saline‐loaded rats at doses of and above 10 mg kg−1, s.c., and 30 mg kg−1, p.o. 8 The prostaglandin‐mediated fall in mean arterial blood pressure elicited in anaesthetized rats by i.v. application of arachidonic acid was not inhibited by prior treatment with EMD 61753 (lO mg kg−1, p.o.). Thus, a blockade of prostaglandin synthesis via inhibition of cyclo‐oxygenase activity does not contribute to the in vivo effects of EMD 61753 and its metabolites. 9 The present experiments therefore indicate that EMD 61753 is a potent, selective and orally‐effective full K‐opioid receptor agonist which has a limited ability to penetrate the blood‐brain barrier and elicit centrally‐mediated sedation, putative aversion, diuresis, and antinociception. The inhibitory actions of systemically‐applied EMD 61753 against hyperalgesic pressure nociception and neurogenic inflammation are mediated peripherally, probably by opioid receptors on the endings of sensory nerve fibres.

Antinociceptive effects of the 5-HT2 antagonist ritanserin in rats: evidence for an activation of descending monoaminergic pathways in the spinal cord.

The antinociceptive properties of the 5-HT2 antagonist ritanserin have been investigated in the writhing test using rats implanted with chronic lumbar catheters. The antinociceptive action of 15 mg/kg ritanserin applied subcutaneously (s.c.) was powerfully inhibited by the intrathecal (i.t.) application of ritanserin (50 nmol), methysergide (15 nmol), yohimbine (20 nmol), alpha-flupenthixol (20 nmol) or naloxone (15 nmol), but not atropine (30 nmol), at doses which themselves produced no change in nociceptive threshold. It is concluded that ritanserin acts supraspinally to activate pain-modulating descending serotonergic, noradrenergic, dopaminergic and possibly opioidergic pathways, while spinopetal cholinergic pathways do not seem to be involved. Hyperalgesic effects of s.c. ritanserin following the i.t. application of methysergide or yohimbine were interpreted in terms of the co-release of an excitatory transmitter, possibly substance P, from descending serotonergic and noradrenergic nerve fibres. The supraspinal mechanism by which ritanserin activates spinopetal pathways and its dependence on 5-HT2 receptors have not yet been established.

Effect of isomazole on the responsiveness to calcium of the contractile elements in skinned cardiac muscle fibres of various species

We report here that the new cardiotonic agent isomazole increases the Ca2+ responsiveness of skinned myocardial fibres from different species. Isomazole (0.01-1 mM) increased the half-maximal, Ca2+-activated force development in atrial fibres from humans and in ventricular fibres from pigs, monkeys, guinea-pigs and rats in a concentration-dependent manner. The maximal force development was also increased in fibres from all species except pig. Isomazole shifted force-calcium curves to the left and enhanced maximal force as demonstrated for guinea-pig and human fibres, indicating increases in both calcium sensitivity and maximum force-generating capability of the contractile apparatus. In guinea-pig tissue, the concentration dependence for the activation of skinned fibres was similar to that for the positive inotropic effect in papillary muscle. These results suggest a direct effect of isomazole on contractile proteins, but the degree to which this effect contributes to the overall positive inotropic action of isomazole remains to be determined.

Avoidance acquisition in adult and senescent rats

Male rats aged 6, 19, or 33 months were trained successively in one- and two-way avoidance tasks. The one-way avoidance test consisted of up to 30 trials given in a single session with the conditional stimulus (CS; 14-kHz tone) presented for either 3 s or for 10 s in separate groups. Senescent rats performed poorest, middle-aged rats intermediately, and young adult rats best. Failure of the longer CS to yield better acquisition than the short CS in the senescent group suggested that the age-related deficit probably did not result from slower responding. In subsequent shuttle box training there was no appreciable age difference in achieving the learning criterion. A compound visual-auditory CS was used, and in further evaluation of well-trained rats it was found that the auditory component was much more effective than the visual component in eliciting avoidance. However, this differential effect of the two stimuli was much weaker in the senescent group than in the young adult group. Nonetheless, these same senescent and adult rats readily learned to make avoidance responses using only the auditory CS, demonstrating that this was an effective stimulus for all age groups.

Central and peripheral actions of the novel κ-opioid receptor agonist, EMD 60400

1 The pharmacological characteristics of the κ‐opioid receptor agonist, EMD 60400, have been investigated, with particular reference to its central and peripheral sites of action and its ability to influence nociception. The κ agonists ICI 197067 and ICI 204448 were tested for purposes of comparison. 2 EMD 60400 and ICI 197067 bind with high affinity (IC50 values of 2.8 and 1.5 nm, respectively) and high selectivity to κ‐opioid receptors. ICI 204448 has a lower binding affinity (IC50 13.0 nm) and selectivity for κ‐opioid receptors. 3 EMD 60400, ICI 197067, and ICI 204448 are full and potent agonists in the rabbit vas deferens in vitro assay for κ‐opioid receptors (IC50 values of 41.8, 15.7 and 15 nm, respectively). 4 Ex vivo binding experiments in mice revealed that EMD 60400 and ICI 197067 were well taken up after s.c. administration. Brain levels of EMD 60400 were lower than those of ICI 197067 at comparable doses, indicating that EMD 60400 does not penetrate into the CNS as well as ICI 197067. 5 Haloperidol‐induced DOPA accumulation in the nucleus accumbens of the rat was dose‐dependently reversed by s.c. application of EMD 60400 and ICI 197067 at doses of and above 3 and 0.3 mg kg−1, respectively. ICI 204448 had no effect on DOPA accumulation at 30 mg kg−1, s.c. 6 Prolongation of hexobarbitone‐induced sleeping time in mice and motor impairment in the rat rotarod test were observed for EMD 60400 at doses above 3 and 2.5 mg kg−1, s.c., respectively, and for ICI 197067 at doses above 0.3 and 0.25 mg kg−1, s.c., respectively. ICI 204448 was inactive in these tests at doses of 30 and 100 mg kg−1, s.c., respectively. 7 EMD 60400 applied s.c. produced dose‐dependent naloxone‐reversible antinociception in the mouse formalin test (1st and 2nd phase ID50 0.44 and 0.47 mg kg−1, respectively) and rodent writhing test (ID50 mouse 0.55 mg kg−1 and rat 0.3 mg kg−1). Furthermore, EMD 60400 was considerably more potent in the rat pressure pain test after the induction of inflammation with carrageenin than under normalgesic conditions (ID50 values 0.1 μg kg−1 and 4.0 mg kg−1, s.c., respectively). The action of EMD 60400 (50 μg kg−1, s.c.) in the hyperalgesic pressure pain test was completely antagonized by injection of the κ‐opioid antagonist, norbinaltorphimine (100 μg) into the inflamed tissue, thus demonstrating the peripheral opioid nature of this effect. 8 EMD 60400 produced dose‐dependent inhibition of neurogenic plasma extravasation elicited by antidromic electrical stimulation of the rat saphenous nerve (ID50 value 0.3 mg kg−1, i.v.). This inhibition was completely antagonized by intraplantar injection of norbinaltorphimine (50 μg). 9 EMD 60400, ICI 197067, and ICI 204448 have diuretic effects in rats at doses of and above 0.1, 0.01, and 0.3 mg kg−1, s.c., respectively. An antidiuretic action was also observed with ICI 197067 at very low doses (3 and 6 μg kg−1, s.c.). 10 Pharmacological and biochemical data therefore indicate that the three κ‐opioid receptor agonists tested here have different tendencies to elicit centrally‐mediated sedation and putative aversion (ICI 197067 > EMD 60400 > ICI 204448) which correspond to their ability to cross the blood‐brain barrier. EMD 60400 combines high affinity and selectivity for the κ receptor with a degree of peripheral selectivity. The peripheral actions of systemically‐applied EMD 60400 against hyperalgesic pressure pain and neurogenic inflammation are very probably mediated by opioid receptors on the endings of sensory nerve fibres.

Antagonistic effects of bisoprolol on several β-adrenoceptor-mediated actions in anaesthetized cats

The beta-adrenoceptor antagonistic activity of i.v. administered bisoprolol ((+/-)-1-[4-(2-isopropoxyethoxymethyl)-phenoxy] -3-isopropylamino-2-propanol, hemifumarate) was studied under two different sets of experimental conditions in anaesthetized cats and compared to the activity of atenolol and propranolol. The responses of several target organs to beta 2-adrenoceptor stimulation were used: inhibition of isoprenaline effects on diastolic blood pressure, hindlimb perfusion pressure, soleus muscle contractility and histamine aerosol-induced bronchoconstriction. The inhibition of isoprenaline-induced tachycardia served as indicator of beta 1-antagonism. The slopes of agonist dose ratio vs. antagonist dose effect were close to unity for propranolol but deviated from unity for atenolol and even more so for bisoprolol. In spite of the ensuing difficulty of comparisons, bisoprolol showed the most pronounced selectivity indices (10-20), followed by atenolol (1-7.5) and propranolol (0.3-1.6). Thus, bisoprolol exhibited a higher degree of beta 1-selectivity in the cat than did atenolol, regardless of the parameter used for measurement of beta 2-antagonism. Propranolol proved to be non-selective or even had a somewhat higher affinity for beta 2- than for beta 1-adrenoceptors.