Makoto Muramatsu

NE-100, a novel sigma receptor ligand: In vivo tests

It has been suggested that sigma receptor antagonists may be useful as antipsychotic drugs. N, N-dipropyl-2-[4-methoxy-3-(2-phenylethoxy)phenyl]-ethylamine monohydrochloride (NE-100) is a novel compound with high affinity for the sigma receptor (IC50 = 4.16 nM), but low affinity (IC50 > 10,000 nM) for D1, D2, 5-HT1A, 5-HT2 and phencyclidine (PCP) receptors. The head-weaving behavior induced by either (+)SKF10047 or PCP was dose-dependently antagonized by NE-100 with oral ED50 at 0.27 and 0.12 mg/kg, respectively. NE-100 did not affect dopamine agonists-induced stereotyped behavior and/or hyperactivity. NE-100 failed to induce catalepsy in rats. These findings indicate that NE-100 may have antipsychotic activity without the liability of motor side effects typical of neuroleptics.

NE-100, a novel potent σ ligand, preferentially binds to σ1 binding sites in guinea pig brain

The selectivity of N, N-dipropyl-2-[4-methoxy-3-(2-phenylethoxy)phenyl]-ehthylamine monohydrochloride (NE-100) for σ1 and σ2 binding sites was studied by means of binding of [3H](+)-pentazocine and [3H]1,3-di-o-tolylguanidine ([3H]HTG) in guinea pig brain. NE-100 inhibited [3H](+)-pentazocine binding to σ1 binding sites potently with an IC50 value of 1.54±0.26 nM while it had a weak effect on [3H]DTG binding to σ2 binding sites. The inhibitory effect of NE-100 on [3H](+)-pentazocine was 55 times more potent than that on [3H]DTG binding. These results suggest that NE-100 is a potent and selective ligand for σ1 binding sites.

Serotonin-2 receptor-mediated regulation of release of acetylcholine by minaprine in cholinergic nerve terminal of hippocampus of rat

5-Hydroxytryptamine (5-HT) inhibited the K+-induced release of [3H]acetylcholine [( 3H]ACh) from slices of the hippocampus of the rat, dose-dependently. Minaprine (3-(2-morpholinoethylamino)-4-methyl-6-phenylpyridazine, Fig. 1) had no effect on the release of [3H]ACh. However, it inhibited the (formula; see text) Fig. 1. Chemical structure of minaprine dihydrochloride. attenuation of the release of [3H]ACh by 5-HT dose-dependently. The 5-HT2 receptor antagonists, mianserine, methysergide and spiperone, prevented the inhibitory effect of the 5-HT, as well as did minaprine. The attenuating effect of 5-HT was not mimicked by the 5-HT1A receptor agonist 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT) and was not prevented by a 5-HT1A and 5-HT1B mixed receptor antagonist, propranolol, or by the 5-HT3 receptor antagonists, cocaine and metoclopramide. Minaprine inhibited the bindings of [3H]5-HT, [3H]8-OH-DPAT and [3H]ketanserin in the hippocampus. The inhibitory effect of minaprine on the binding of [3H]ketanserin was more marked than on the binding of [3H]5-HT and [3H]8-OH-DPAT, and was non-competitive. The Ki value of minaprine for the binding of [3H]ketanserin was 2.9 microM. The inhibitory effect of 5-HT on the release of [3H]ACh was observed in the presence of tetrodotoxin. By electrolytic lesioning of the medial septum, the K+-induced release of [3H]ACh from the slices of hippocampus was significantly reduced and the release was no longer inhibited by 5-HT. The lesioning significantly decreased the binding of [3H]ketanserin in the hippocampus, with hardly any reduction in the binding of [3H]5-HT and [3H]8-OH-DPAT.(ABSTRACT TRUNCATED AT 250 WORDS)

5-HT2 antagonists and minaprine block the 5-HT-induced inhibition of dopamine release from rat brain striatal slices

5-Hydroxytryptamine (5-HT) inhibited the K+-induced [3H]dopamine [( 3H]DA) release from slices of rat striatum. Minaprine (3-(2-morpholinoethylamino)-4-methyl-6-phenylpyridazine) attenuated the inhibitory effect of 5-HT in a dose-dependent manner. 5-HT2 receptor antagonists, ketanserin and mianserin, prevented the effect of 5-HT as well as minaprine did. The inhibitory effect of 5-HT was not mimicked by a 5-HT1A receptor agonist, 8-hydroxy-2-(di-n-propylamino)tetralin (8-OH-DPAT), and was not prevented by a 5-HT1A and 5-HT1B mixed receptor antagonist, propranolol. Minaprine was a potent inhibitor of the binding of [3H]ketanserin to binding sites in the striatum over the concentration range 10(-6)-10(-4) M. Lesion of the medial forebrain bundle with 6-hydroxydopamine (6-OHDA) significantly reduced the K+-induced [3H]DA release from the striatum and release was no longer inhibited by 5-HT. Lesioning, however, did not change significantly the [3H]ketanserin binding in the striatum. These results suggest that minaprine suppresses the inhibitory effect of 5-HT on DA release in the striatum via the inhibition of 5-HT binding at the 5-HT2 receptor on the nerve terminal of the DA-ergic neuron and, further, that the proportion of the 5-HT2 receptor site which is located on the nerve terminal of the DA-ergic neuron is small in the striatum.

Inhibition of Gastric H+,K+-ATPase by Flavonoids: A Structure-Activity Study

Gastric H+, K(+)-ATPase plays a pivotal role in the final step of gastric acid secretion. Over 80 flavonoids, including flavones, flavanones, isoflavones and anthocyanidins were examined for their in vitro effect on gastric H+, K(+)-ATPase and some were found to be inhibitors of this enzyme. Kinetic studies showed that the inhibition of H+, K(+)-ATPase by flavonoids was competitive with respect to ATP, and non-competitive with respect to K+. Structure-activity analysis revealed the following: (1) The inhibitory potency of flavonoids depends on the number of hydroxyl groups up to four per molecule and that above this, no marked enhancement is seen; (2) The hydroxylation pattern is an important determinant of inhibitory potency. Two adjacent hydroxyl groups (catechol-type), three adjacent hydroxyl groups (pyrogallol-type) or hydroxyl groups at C-3, C-5 and C-7 are a minimum requirement for high potency inhibition; (3) Protection of the hydroxyl group(s) by glycosylation or methylation decreases potency; (4) Saturation of the C-2-C-3 double bond results in a decrease in potency; and (5) A ketone at C-4 is not essential for inhibition.

Inhibition of gastric H+, K+-ATPase by chalcone derivatives, xanthoangelol and 4-hydroxyderricin, from Angelica keiskei Koidzumi

Abstract— Two chalcone derivatives, xanthoangelol (I) and 4‐hydroxyderricin (II) isolated from Angelica keiskei Koidzumi, inhibited pig gastric H+, K+ ‐ATPase with IC50 values of 1.8 and 3.3 μM, respectively. The inhibition by I or II was competitive with respect to ATP and was non‐competitive with respect to K+. I and II also inhibited K+, stimulated p‐nitrophenyl phosphatase, with IC50 values of 1.3 and 3.5 μM, respectively. Proton transport in‐vitro was inhibited by I or II, in a dose‐dependent manner. I at 100 mg kg−1, i.p. significantly inhibited acid secretion and the formation of stress‐induced gastric lesions. These results suggest that the antisecretory effect of I is due to the inhibition of gastric H+, K+‐ATPase.

Gastric H+, K+-ATPase inhibition by catechins

Abstract— Five catechins, (+)‐catechin, (–)‐epicatechin, (–)‐epicatechin gallate, (–)‐epigallocatechin and (–)‐epigallocatechin gallate, inhibited gastric H+, K+‐ATPase activity with IC50 values ranging from 1·7 × 10−4 to 6·9× 10−8 m, with (–)‐epigallocatechin gallate as the most potent inhibitor. The intensity of inhibitor activity paralleled the number of phenolic hydroxy groups in the molecule. The inhibition of the enzyme by (–)‐epicatechin was competitive with respect to ATP and noncompetitive with respect to K+. These findings suggest that the anti‐secretory and antiulcerogenic effects of catechins previously reported, are due to their inhibitory activity on gastric H+, K+ ‐ATPase.

Inhibition of gastric H+,K+-ATPase and acid secretion by cassigarol A, a polyphenol from Cassia garrettiana Craib

The effects of cassigarol A, a naturally occurring polyphenol, on gastric H+,K(+)-ATPase and gastric acid secretion were studied. Cassigarol A inhibited H+,K(+)-ATPase and K-stimulated p-nitrophenyl phosphatase from hog gastric mucosa with 50% inhibition of 1.2 x 10(-6) and 6.3 x 10(-6) M, respectively. The kinetic study showed that the inhibition of H+,K(+)-ATPase by cassigarol A was competitive with respect to ATP and non-competitive with respect to K+. Cassigarol A inhibited both H+,K(+)-ATPase-mediated proton transport and 2-deoxy-D-glucose-induced acid secretion. On the other hand, cassigarol A acetate, in which phenolic hydroxy groups are acetylated, was not effective in the inhibition of enzyme activity and acid secretion. These results indicate that cassigarol A is a potent inhibitor of gastric H+,K(+)-ATPase, that the anti-secretory activity of cassigarol A is related to the inhibition of H+,K(+)-ATPase and that an important moiety of cassigarol A in the interaction with the enzyme is the phenolic hydroxy groups.

Effects of restraint and water-immersion stress and insulin on gastric acid secretion in rats

Effects of restraint and water-immersion stress (RWIS) and of insulin injection on gastric acid secretion were investigated in relation to blood glucose levels and to brain glucose uptake in rats. Venous blood glucose levels (VBG) were significantly increased while arterial blood glucose level (ABG) was slightly increased by RWIS. In contrast, ABG and VBG were significantly decreased by administration of insulin; the decrease in ABG was greater than that in VBG. Glucose uptake into the brain, calculated from the ABG-VBG, was significantly decreased both by RWIS loading and by insulin administration. The uptake of [14C] 2-deoxy-D-glucose [( 14C]-2DG) into the brain was also significantly decreased in RWIS-loaded or insulin-treated rats. Gastric acid output was significantly increased both by RWIS loading and by insulin administration. The increased acid output paralleled the decrease of glucose uptake into the brain in RWIS-loaded and insulin-treated rats. Results suggest that RWIS-induced gastric acid secretion is regulated by brain glucose uptake and that this gastric acid secretion is a hypothalamic neuron-mediated event as is insulin-stimulated gastric acid secretion.

Characteristics of binding of [3H]NE-100, a novel sigma-receptor ligand, to guinea-pig brain membranes

We examined the characteristics of binding of radiolabeled N,N-dipropyl-2-[4-methoxy-3-(2-phenylethoxy)phenyl]-ethylamine monohydrochloride ([3H] NE-100), a highly potent and selective sigma-receptor ligand, to guinea-pig brain membranes. [3H]NE-100 showed saturable and reversible binding to sigma binding sites. A dissociation constant (Kd) and maximal numbers of binding sites (Bmax) obtained from Scatchard plot analysis were 1.2 ± 0.1 nM and 1049.3 ± 115.1 fmol/mg protein (n = 3), respectively. NE-100 was the most potent inhibitor of [3H]NE-100 binding among several structurally dissimilar sigma-receptor ligands, including haloperidol and ( + )-pentazocine. ( + )-Benzomorphanes had more than a 10-fold potent inhibitory activity over (−)-benzomorphanes, with regard to [3H]NE-100 binding. The binding of [3H]NE-100 was not influenced by histaminergic, dopaminergic, adrenergic, serotonergic cholinergic or glutaminergic agents at 10−7 M. GTP-γ-S and phenytoin also did not affect the binding of [3H]NE-100. A higher [3H]NE-100 binding was observed in the cerbellum and medulla oblongata. Except for the nuclear fraction, the highest level of [3H]NE-100 binding to subcellular fractions was observed in microsomal fractions. These results suggest that NE-100 selectively binds, with a high affinity, to sigma-1 binding sites in guinea-pig brain membranes, as an “antagonist”.