Vito Margotta

A pharmacological study of cocaine activity in planaria.

Planaria has been proposed as a suitable research model in neurobiology because of its relatively simple organization. Dopaminergic agonists induce in this flatworm typical hyperkinesias that can be antagonized by dopaminergic blocking agents. The neurochemical basis of the effects of cocaine in vertebrates has not been fully elucidated, but the inhibition of catecholamine reuptake at a presynaptic level seems to play an important role. In this study we analyzed the involvement of the dopaminergic system in the mechanism of action of cocaine in planaria. The dose-related effects of cocaine on planaria motility and the response to cocaine treatment associated with the administration of specific D1 or D2 dopamine agonists and antagonists were investigated. The effects of reuptake inhibitors on cocaine activity were also studied. Planaria specimens treated with low doses of cocaine become motionless, whereas high doses induce a typical behavioural response, identical to the response induced by specific D2 agonists. This response is inhibited by a D2 selective blocking agent. Nomifensine, a specific dopamine reuptake inhibitor, induces a mixed D1/D2 response. The results of these experiments are discussed, also in relation with the conservation of dopaminergic receptors during evolution.

Proposal of a New Model with Dopaminergic-Cholinergic Interactions for Neuropharmacological Investigations

The motor system of Dugesia gonocephala shows a striking similarity with the extrapyramidal system of high vertebrates and of man with the evidence of correlations between dopaminergic and cholinergic neurons. The utilization of this model seems to be useful in testing drugs which presumably act on dopaminergic or cholinergic transmission. In this model, the quantification of animal behaviour seems considerably easier when compared with the difficulties met in other animal models commonly employed. Besides, it might be anticipated that this model, if correctly used, can display interesting perspectives also in neuroendocrinological investigations.

Radioimmunological and immunocytochemical demonstration of Met-enkephalin in planaria.

1. An anti-Met-enkephalin serum has been used for radioimmunological and immunocyto-chemical demonstration of Met-enkephalin in planaria. 2. The serum showed a high degree of sensitivity and specificity. 3. Radioimmunoassay demonstrated in planaria extracts a considerable concentration of Met-enkephalin. 4. Met-enkephalin was localized by immunocytochemical methods in neurons and neuropil.

Opioid-dopamine interaction in planaria: a behavioral study.

The behavioral response of planaria to the exposure to selective opioid agonists was studied. The mu agonist [d-ala2, N-methyl-Phe4,Gly5-ol]enkephalin (DAMGO) and the 6 agonist [D-Pen2, D-Pen5]enkephalin (DPDPE) failed to alter motor activity at all doses tested. Low doses of the selective kappa agonist (+/-)-trans-U-50-trans-3,4-dichloro-N-methyl-N[2-(1-pyrrodinyl)-cyclohexyl]benzene acetamide methasulphonate (U50, 488) and bremazocine-HCl increased motor activity leading to C-like position (CLP) and screw-like hyperkinesia (SLH). These changes were identical to those seen previously with the exposure to D2 or D1 dopamine receptor agonists, respectively. Higher doses of kappa agonists produced the enhancement of CLP and SLH together with robust snake-like movements (SLM). This latter response, that was typical of stimulation of kappa opioid receptors, was blocked by co-exposure to naloxone or the selective kappa antagonist Nor-binaltorphimine (Nor-BNI). Finally, co-exposure to sulpiride or SH-23390 respectively blocked the CLP or SLH response produced by U50,488 or bremazocine. Our data indicate the presence of kappa opioid receptors in planaria and suggest the functional interaction between the opioid and dopamine system in this simple animal model.

Aminergic neurotransmitters and adenylate cyclase in hydra

Serotonin, dopamine and, in lesser amounts, norepinephrine were detected in Chlorohydra viridissima with electrochemical detection coupled to liquid chromatography (LCED). Treatment with reserpine induces a significant decrease in amine levels. Adenylate cyclase was found in Hydra tissue; the enzyme is stimulated by Mg, Mn and F and sensitive to guanine nucleotide activation. Dopamine, serotonin, GSH and glutamate do not affect cyclase activity.

Cannabinoid-induced stimulation of motor activity in planaria through an opioid receptor-mediated mechanism

Planaria, the most primitive example of centralization and cephalization of the nervous system along phylogeny, shows specific stereotyped behavioral patterns following exposure to drugs acting on neural transmission. In this study, the authors investigated the effects of exposure to the synthetic cannabinoid receptor agonist WTN55212.2 on motor activity in planaria. WTN55212.2 produced dose-dependent stimulation of motor behavior. High doses of the drug caused stereotyped activities identical to those seen previously with opioid agonists. These effects were antagonized by coexposure to cannabinoid or opioid receptor antagonists. The results indicate that functional interactions between cannabinoid and opioid systems are highly conserved along phylogeny, at least at the behavioral level.

Naloxone enhances cAMP levels in planaria

Abstract 1. In planaria ( Dugesia gonocephala s.l. ) treatment with dopaminergic agonists induces screw-like hyperkinesias and a rise in cAMP levels. 2. Treatment with dopaminergic antagonists causes a decrease in motor activity and cAMP levels. 3. Morphine treatment induces a reduction of motor activity and a decrease in cAMP levels. 4. Naloxone induces strong hyperkinesias and a significant rise in cAMP. Pretreatment with reserpine or with haloperidol inhibits naloxone action. 5. A model similar to the one operative in the nigro-striatal system of vertebrates is proposed for planaria.

The pigmentary system of planaria

SummaryThe pigmentary system of the planaria, Dugesia gonocephala s.l. (Platyhelminthes, Turbellaria, Tricladida), has been studied by light and electron microscopy. The system consists of granules contained in chromatophore-like cells embedded in the parenchyma. The cell processes penetrate between the muscle layers and extend to the sub-epidermal basal lamina. The nature of the pigment and the comparative anatomical significance of the chromatophore structure is discussed.

PCNA positivity in the mesencephalic matrix areas in the adult of a Teleost, Carassius carassius L.

In normal adult Teleostei, classical histology and autoradiographic techniques have been used to identify the telencephalic hemispheres on the dorso-lateral (zona germinativa dorsalis) and medial (zona germinativa ventralis) surfaces in the ependymal and/or peri-ependymal zone as elements capable of proliferating. In inequivocally adultCarassius carassius specimens we revisited these «matrix areas» using a recent proliferative marker that detects the expression of the Proliferating Cellular Nuclear Antigen (PCNA), a reliable immunocytochemical test of the essential role played by PCNA in the cell cycle, its ubiquity and the theoretical bases on which this method rested. It was found that in the telencephalon the distribution of the DNA synthesis activity is not continuous and uniform in an antero-posterior direction. A non negligible PCNA positivity is actually detectable at the anterior pole of the hemispheres and above all posteriorly, at the level of thezona germinativa ventralis while undifferentiated elements are visible in thezona germinativa dorsalis, in very small numbers in the lateral portion and sporadically in the upper section. Overall it would seem that in theCarassius telencephalon spontaneous neurogenic potential has now become residual. These findings are discussed in the light of literature data and above all of the age of our specimens.RiassuntoNei Teleostei adulti normali, l’istologia classica e le tecniche autoradiografiche, avevano identificato negli emisferi telencefalici sulle superfici dorso-laterale (zona germinativa dorsalis) e mediale (zona germinativa ventralis), in sede ependimale e/o peri-ependimale, elementi in grado di proliferare. InCarassius carassius inequivocabilmente adulti abbiamo rivisitato tali «aree matrici», utilizzando un recentemarker proliferativo che rivela l’espressione dellaProliferating Cellular Nuclear Antigen (PCNA), test immunocitochimico affidabile per il ruolo essenziale svolto dalla PCNA nel ciclo cellulare, la sua ubiquitarietà e le basi teoriche su cui poggia tale metodica. Si è rilevato che nel telencefalo la distribuzione dell’attività di sintesi del DNA non è, in senso antero-posteriore, continua ed uniforme. Infatti, una non trascurabile PCNA positività è evidenziabile al polo anteriore degli emisferi, specie posteriormente a livello dellazona germinativa ventralis, mentre elementi indifferenziati si osservano nellazona germinativa dorsalis con molta parsimonia nella porzione laterale ed episodicamente nel tratto superiore. In una visione d’insieme sembrerebbe che nel telencefalo delCarassius le potenzialità neurogeniche spontanee siano ormai residuali. Tali rilevanze vengono discusse alla luce dei dati forniti dalla bibliografia e soprattutto dell’età dei nostri esemplari.

PCNA positivity in the telencephalic matrix areas in the adult of a frog,Rana esculenta

Using a recent immunocytochemical protein-type marker having an auxiliary function of DNA polymerase delta which is expressed during the DNA synthesis phase of the cell cycle, namely theProliferating Cellular Nuclear Antigen (PCNA), we ascertained that the anti-PCNA labelling locations allow several regions of the telencephalon ofRana esculenta normal adults to be identified. The immunocytochemical aspects are the same as those previously found in the telencephalon of adult Anurans, either normal or subjected to a wide range of experimental situations, using both classical histology and autoradiographic methods. The survival in the adult of undifferentiated elements observed by us in thezona germinativa dorsalis and in thezona germinativa ventralis is examined in the light of literature evidence. The extent and possible role of these elements as the expression of continual spontaneous neurogenic episodes, as well as more substantial and widespread neurogenic events due to natural or experimental causes are discussed.RiassuntoAvvalendoci di un recentemarker, di tipo immunocitochimico, di natura proteica e con funzione ausiliaria della DNA polimerasi delta, il quale è espresso, nel ciclo cellulare, durante la fase di sintesi del DNA, laProliferating Cellular Nuclear Antigen (PCNA), abbiamo evidenziato che la dislocazione delle marcature anti-PCNA permette di identificare nel telencefalo di adulti normali diRana esculenta alcune regioni. Gli aspetti immunocitochimici sono sovrapponibili a quanto già evidenziato nel telencefalo di Anuri adulti, sia normali che sottoposti a svariate situazioni sperimentali, mediante l’istologia classica e le metodiche autoradiografiche. Alla luce dei dati forniti dalla letteratura, sulla sopravvivenza nell’adulto di elementi indifferenziati da noi osservati nellazona germinativa dorsalis e nellazona germinativa ventralis, viene discussa l’entità ed il loro eventuale ruolo quale espressione di continui episodi neurogenici spontanei, come pure di eventi neurogenici più consistenti e diffusi, indotti da cause naturali o sperimentali.