Effects of methyl-eugenol administration on behavioral models related to depression and anxiety, in rats.

Croton zehntneri (Cz) is a popular plant in Brazilian folk medicine. Recently, the use of its essential oil showed depressive activity response in the central nervous system (CNS). Chemical studies show that the main compound of this oil is the methyl-eugenol (ME). This work seeks to evaluate the ME activity in behavioral models of depression and anxiety, in the rat. Male rats (60 days old) were divided into four groups (n = 10) and treated with doses of 1.0, 3.0 and 10.0 ml/100 g body wt., v.o., of ME (experimental) and saline (control). One hour after treatment, they were observed in the forced swimming test and 15 min later in the open-field test. A decrease was observed in the immobility time during the forced swimming test for all experimental groups, in comparison with control group (C = 168.8 +/- 27.3; 1.0 microl = 139.1 +/- 23.5; 3.0 microl = 137.2 +/- 18.7 and 10.0 microl = 139.8 +/- 23.6). The open-field results showed no differences in comparison to the control group. The same was observed for social interaction, plus-maze and holeboard tests, suggesting no alterations in anxiety behavior. These data suggest that ME administration induced antidepressive CNS alterations, expressed by the smallest immobility in the swimming model, and not of a level able to alter motor and exploratory activity in the open-field. The absence of effects observed in the open-field can be a result of the experimental contingency, taking low anxiety levels. These data are in contradiction to observations with Cz essential oil in these models.

Okadaic acid, useful tool for studying cellular processes.

One of the most interesting groups of substances of marine origin, from structural and pharmacological points of view are polyether toxins, which generally present a great diversity in size and potent biological activities. The subject of this review is limited to okadaic acid (OA). It was the first example of a group of polyether toxins produced by marine microalgae, which is responsible for the natural phenomena known as Diarrhetic Shellfish Poisoning, DSP red tides. These toxins are accumulated in the digestive glands of the shellfish with a disastrous effect upon the shellfish industry in many parts of the world. Thus, it has been demonstrated that OA is a highly selective inhibitor of protein phosphatases type 1 (PP1) and 2A (PP2A), subsequently that it causes dramatic increases in phosphorylation of numerous proteins as well as being a potent tumour promoter. For that reason, OA is an extremely useful tool for studying the cellular processes that are regulated by reversible phosphorylation of proteins as signal transduction, cell division and memory.

Isolation and structural determination of DTX-6, a new okadaic acid derivative.

Diarrhetic shellfish poisoning is an illness caused by toxins accumulated in shellfish and produced by dinoflagellates, mainly of the Dinophysis and Prorocentrum genera. This paper reports the isolation and spectroscopic structural elucidation of a new compound, DTX-6 (2), an ester derivative of okadaic acid (OA) (1), isolated from an artificial culture of strain PLV2 of Prorocentrum lima.

Influence of amino acids on okadaic acid production.

Okadaic acid (OA) (1)) was the first example of a group of polyether toxins known to be produced by marine microalgae, which are responsible for the natural phenomena known as Diarrhetic Shellfish Poisoning (DSP) red tides. It is also a highly selective inhibitor of protein phosphatases type 1 (PP1) and 2A (PP2A), as well as being a potent tumour promoter. For these reasons, OA is an extremely useful tool for studying cellular processes and an important standard for polluted shellfish control. In this paper, we report on a double objective: to improve the production of toxins and verify the apparent participation of amino acids in the formation of these polyethers by monitoring their influence on the promotion of growth, total cell yield and increased in toxicity in Prorocentrum lima of the PL2V strain in batch cultures, in a modified K medium.

Evaluation of the cytotoxic activity of polyethers isolated from Laurencia.

In this paper, we report on the conformational analysis of several polyether triterpenes with a squalene carbon skeleton which exhibited significant cytotoxic activity using a Monte Carlo conformational search and spectroscopical data. These studies indicate that the conformation of the side chain C-14/C-19 and the arrangement and direction of this chain may be among the fundamental factors related to the activity of this type of metabolites.

Complexation of okadaic acid: a preliminary study.

A complexation study was carried out with okadaic acid (OA) and the univalent metal ions Li+, Na+ and K+, and the divalent metal ions Ca2+ and Mg2+. K+ binding was observed identical with a complex obtained from the natural source (OAC). The pharmacological trials demonstrated that this cation has a very important influence on the pharmacological activity of okadaic acid.

An analysis of the mechanisms involved in the okadaic acid-induced contraction of the estrogen-primed rat uterus.

The contractile effect of okadaic acid (OA) and its derivatives was investigated in the rat uterus. OA (20 microM) induced a transient contraction which, after plateauing, slowly decreased. The structurally related compound okadanol (20 microM) failed to induce any significant contraction. Conversely, the synthetic compound methyl okadaate (20 microM) and the naturally occurring ester 7'-hydroxy-4'-methyl-2'-methylen-hept-4'(E)-enyl okadaate (20 microM) were as active as the free acid. The OA-induced contraction was unaffected in the presence of neomycin (5 mM), mepacrine (30 microM), 1-[N,O-bis(1,5-isoquinolinesulfonyl)-N-methyl-L-tyrosyl]-4-phenylpiperaz ine (10 microM), calphostin C (3 microM) and 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (30 microM). The calmodulin inhibitor N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride (100 microM) did not modify the amplitude of the OA-induced contraction but significantly increased the rate of tension decay. The myosin light chain kinase inhibitor 1-(5-chloronaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine hydrochloride (1 mM) significantly reduced the peak amplitude of the contraction. Staurosporine (0.03-0.1 microM) did not modify the contractile component of the OA-induced response but inhibited the subsequent decrease in tension. In freshly dispersed myometral cells loaded with the fluorescent Ca++ indicator indo 1, OA did not produce any significant increase in [Ca++]i. OA (5- to 90-min contact) also failed to modify the intracellular levels of arachidonic acid, compared with basal values. These data suggest that in the rat uterus 1) the contractile effect of OA (20 microM) is specifically mediated by inhibition of protein phosphatases type 1 and/or 2A and is related to a direct interaction with the contractile machinery; 2) the decreasing phase of the OA-induced mechanical response could be mediated by a staurosporine-sensitive protein kinase different from protein kinase C.

Inhibitory effects of okadaic acid on rat uterine contractile responses to different spasmogens.

In the present study, we examined the effects of okadaic acid, a selective inhibitor of type 1 and 2A protein phosphatases, on the mechanical responses evoked by oxytocin, K(+)- and Na(+)-modified solutions and ouabain in estrogen-primed rat myometrium. Oxytocin elicited a rapid, phasic contraction followed by rhythmic oscillations. The phasic response was partially resistant to the absence of external Ca2+. Okadaic acid (1 microM) and the L-type calcium channel blocker nifedipine (1 microM) abolished the oscillatory component and reduced the initial, phasic response to about 80% of the control response. High K+ (60 mM) solution, ouabain (1 mM), K(+)-free medium and low Na+ (25 mM) solution induced extracellular Ca(2+)-dependent biphasic responses composed by an early rapid (KCl, ouabain and K(+)-free solution) or slower developed (25 mM Na+ solution) phasic contraction followed by a sustained increase in tension. Okadaic acid and nifedipine, alone or in combination, abolished or decreased similarly the contractile response evoked by these stimulants. The okadaic acid- and nifedipine-insensitive responses to ouabain, K(+)-free and low Na+ solution were enhanced by increasing the extracellular concentration of Ca2+ in the medium and were inhibited in a dose-dependent manner by amiloride (0.05-0.5 mM). These data suggest that, in estrogen-primed rat uterus, dephosphorylating mechanisms by OA-sensitive protein phosphatases play an important role in regulating myometrial contractions elicited by Ca2+ entry through voltage-sensitive Ca2+ channels.

Calcium dependence of the mechanical response evoked by okadaic acid in smooth muscle.

The effects of okadaic acid (OA), obtained from a culture of the marine dinoflagellate Prorocentrum lima were studied on isolated strips of rat myometrium. The contractile response evoked by OA at 5, 10, and 20 microM in normal physiological solution was unaffected in the presence of tetrodotoxin (10 microM), indomethacin (3 microM), or a cocktail of antagonists which blocked muscarinic, adrenergic, histaminergic, serotonergic, and opioid receptors. Similarly, the response to OA was unaffected in the presence of nifedipine at a concentration (1 microM) which completely or highly blocked the response to KCl (60 mM), oxytocin (1 microM), or acetylcholine (100 microM). In a Ca(2+)-free 1 mM EGTA-containing solution, the response to 10 and 20 microM OA was slightly but significantly reduced whereas the response to 5 microM OA was abolished. However, a response similar to that evoked in Ca(2+)-containing solution was observed when 5 microM OA was added to the bath in the presence of 1 microM oxytocin or 160 microM vanadate in a Ca(2+)-depleted solution with 1 mM EGTA. These data suggest that the response of rat myometrium to OA (> or = 5 microM) is not mediated through activation of membrane receptors or neurotransmitter release nor by cyclo-oxygenase products. The response to OA (10 and 20 microM) is highly resistant to the absence of calcium in the medium and does not seem to involve calcium entry through dihydropyridine-sensitive Ca2+ channels.(ABSTRACT TRUNCATED AT 250 WORDS)

Influence of calcium on the effects of okadaic acid and its interaction with caffeine and theophylline in rat myometrium.

The effects of okadaic acid (OA), a monocarboxylic acid produced by marine dinoflagellates belonging to the genera Dinophysis and Prorocentrum, and their interactions with theophylline and caffeine were studied on the rat-isolated uterus in a calcium-containing medium and a calcium-free medium in the presence of 10(-3) M EGTA. Okadaic acid (5 x 10(-6) to 5 x 10(-5) M) induced a concentration-dependent contraction of the rat-isolated uterus corresponding, with 5 x 10(-5) M, to 142.3 +/- 6.1% (n = 7) of the contraction induced by oxytocin 10(-6) M. The time to peak tension was inversely proportional to the maximum effect produced. The contraction was not sustained and was followed by a concentration-dependent decrease in tone. In a Ca(2+)-free medium containing 10(-3) M EGTA the contractile effects of OA were significantly inhibited or reduced. A 30 min pretreatment with theophylline (3 x 10(-3) M) or caffeine (2 x 10(-2) M) significantly reduced, in a Ca(2+)-containing medium, the maximum contractile effect of OA 10(-5) and/or 2 x 10(-5) M and shortened the relative time to peak tension. In a Ca(2+)-free medium containing 10(-3) M EGTA, only the second effect was observed. With a 1 min pretreatment and in a Ca(2+)-containing medium, theophylline 3 x 10(-3) M and caffeine 10(-2) M did not modify the maximum effect of OA 10(-5) M but shortened the time to peak tension.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of okadaic acid on the human isolated bronchus.

The effects of okadaic acid, a polyether derivative of a 38-carbon monocarboxylic fatty acid obtained from a culture of the marine dinoflagellate, Prorocentrum lima, were studied on the human isolated bronchus. In low concentrations (0.01 and 0.03 microM), okadaic acid had no significant effect of its own on the human isolated bronchus, but in higher concentrations (0.1-10 microM) it induced a series of contractions and relaxations. The first contraction was of low intensity (5% of maximum response to acetylcholine 3 mM) and occurred early. The second contraction had a higher amplitude (30% of maximum response to acetylcholine 3 mM) and reached its peak with okadaic acid 0.3 microM. At higher concentrations (1-10 microM), following a relaxation phase, a later rebound contraction occurred between 70 and 120 min and corresponded to 40% of the maximum response to acetylcholine 3 mM. In addition, okadaic acid inhibited or abolished the contractile response evoked by either KCl 60 mM or acetylcholine 3 mM with IC50 of 0.04 and 0.12 microM, respectively. The second contraction evoked by 0.3 microM okadaic acid was partially inhibited in the presence of the Ca2+ channel blocker, nicardipine 1 microM, or after incubation of the human bronchus in a Ca(2+)-free solution and it was completely abolished in the presence of CdSO4 0.1 mM.(ABSTRACT TRUNCATED AT 250 WORDS)

Inhibitory and contractile effects of okadaic acid on rat uterine muscle.

The effects of okadaic acid and its interactions with various agents known to increase, by different mechanisms, the intracellular levels of cyclic AMP and/or cyclic GMP were investigated in isolated strips of rat myometrium. Okadaic acid showed inhibitory effects at concentrations between 10(-7) M and 3 x 10(-6) M. At higher concentrations, a biphasic, contractile and then relaxant response was observed. The results obtained suggest that, in rat uterine smooth muscle, the inhibitory effects of okadaic acid are not entirely mediated by the activation of cyclic AMP- and/or cyclic GMP-dependent pathways. The data also point to the existence of a clear interaction between okadaic acid and methylxanthines, although further studies are needed to clarify the mechanisms involved in this interaction.