Effects of arachidonoyl dopamine, haloperidol, and their mixtures on regeneration of freshwater Hydra attenuata

Arachidonoyl dopamine and haloperidol, both separately and in different combinations, inhibit regeneration of the gastral and basal regions of hydra. In addition, both substances induce stable anomalies of morphogenesis in the form of outgrowths and additional tentacles in gastral regenerates. In the presence of both substances at different combinations, anomalies either do not appear altogether, or exist for a short time, thus suggesting the normalization of morphogenesis. Possible mechanisms underlying the effects of these substances are discussed.     

Interaction of glutathione analogues with Hydra attenuata gamma-glutamyltransferase.

gamma-Glutamyltransferase activity was studied in extracts of the cnidarian Hydra attenuata. The binding of gamma-glutamyl peptide analogues to the enzyme was studied by observing their effects on heat denaturation and their inhibition of p-nitroaniline release from gamma-glutamyl p-nitroanilide. Neither position-1 analogues, in which the gamma-glutamyl moiety was changed to a beta-aspartyl (beta-Asp-Abu-Gly) or an alpha-glutamyl (Glu-Abu-Gly) linkage, nor glutamate protected the enzyme against inactivation at 58 degrees C. GSH (reduced glutathione), gamma-Glu-Abu-Gly and gamma-Glu-Met on the other hand did prevent heat denaturation. GSH and analogues of GSH were competitive inhibitors of p-nitroaniline release, but those analogues in which glycine was replaced by 2-aminoisobutyrate, phenylalanine, leucine or tyrosine had Ki values that were approximately five times those of analogues with the cysteine residue replaced.     

Modulation of Hydra attenuata rhythmic activity. Photic stimulation.

We investigated in Hydra attenuata the possibility of altering more or less permanently and in different environmental conditions, the frequency of Contraction Pulse Trains (CPT's) associated with the rhythmic spontaneous contraction activity, by repetitive light stimuli of variable duration, frequency and amplitude. The CPT's activity of various pieces of Hydra has been also investigated in indisturbed conditions and under stimulation. The following observations have been performed. 1. A transient effect, consisting of an increase or a decrease of CPT's frequency, occurs respectively after an abrupt decrease or increase of the light level. 2. If Hydra is stimulated by repetitive light pulses of 0.5-10 sec duration, at a frequency different from the CPT's average one, the CPT's frequency modifies; if the stimulation frequency is included in a range not too much up or below that of CPT's the new CPT's frequency equals exactly that of stimulation; close to this range the CPT's frequency is a multiple or submultiple of that of stimulation. 3. No habituation to such repetitive stimulation was found. 4. The phase relation between CPT's at the new frequency and light stimuli is a function of the difference between CPT's and stimulation frequencies. 5. Stimulation with repetition of light and darkness periods of some minutes duration induces activity only or mainly during darkness. 6. Modification of CPT's frequency by means of repetitive light stimulation [of the type mentioned either in 2) or 5)] has been observed also with hypostomal preparations. 7. With cessation of the light stimulation, the new CPT's frequency of the whole animal lasts in darkness for a time (10-85 min) that is about 5-10 times longer than that necessary to obtain CPT's syncronization with stimulation. 8. The influence of the light intensity level on transient CPT's frequency variation (see 1), CPT's inhibition and stimulation, promptness of entrainment, range of entrainability, phase relation between entrained CPT's and stimuli, retention time of entrained rhythm has been examined, together with the influence of the reversal of polarity of light transitions on CPT's inhibition and entrainment.     

Differential binding of concanavalin A by dissociated cells of Hydra attenuata

Summary Living dissociated cells of hydra were exposed to fluorescein- and ferritin-conjugated concanavalin A (con A) and observed by light and electron microscopy. Fluorescence microscopy indicated that the isolated cells bound con A differentially; epidermal battery cells showed the greatest binding, whereas small cells belonging to the interstitial cell class displayed the lowest levels of binding. Mature nematocytes had strong localized con A binding at the opercular region. Electron microscopy permitted accurate identification of interstitial cells, early nematoblasts, and nerve cells. The use of ferritin-labeled con A allowed quantitative assessment of lectin binding on these cells. There were significantly fewer con A-binding sites on interstitial cells as compared to nematoblasts and nerve cells, and the amount of con A binding appeared to increase with the maturation of nematocysts from nematoblasts. The findings are discussed in relation to a likely role of cell surface glycoconjugates in the development of positional signals and intercellular junctions that govern final positioning of nematocytes and nerves in hydra.     

Modulation of Hydra attenuata rhythmic activity: phase response curve.

We investigated the effect of photic stimulation on the frequency of Hydra attenuata column contractions. We used positive or negative abrupt light transitions, single or repetitive light or darkness pulses, and alternation of light and darkness periods. The main results are: (a) The frequency of the contraction pulse trains (CPTs) varies transiently in response to an abrupt variation of the light intensity. (b) CPTs in progress can be inhibited by different types of photic stimuli. (c) The response time to a single photic stimulus varies during the inter-CPT interval and depends also on the polarity of the stimulus. (d) The CPTs are entrainable with repetitive light stimulation of various frequencies. (e) Long-lasting variations of the frequency of CPTs occur after the end of a repetitive light stimulation. We suggest that the mechanism responsible for the rhythym of column contractions is quite similar to that on which other biological rhythmic phenomena are based.     

Functional organization of battery cell complexes in tentacles of Hydra attenuata

Ultrastructural and light microscopic observations on the organization of thick and thin regions of hydra's tentacles, made on serial sections and on whole fixed, plastic-embedded tentacles, reveal the existence of two levels of anatomical order in the tentacle ectoderm: (1) The battery-cell complex (BCC), composed of a single epitheliomuscular cell (EMC) and its content of enclosed nematocytes and neurons; and (2) the battery cell complex ring (BCC ring), an arrangement of 4 or more BCCs into larger units organized as rings around the circumference of the tentacle. All EMCs of the distal tentacle appear to contain batteries of nematocytes, and are, therefore, called “battery cells.” Apart from battery cell complexes and migrating nematocytes, there are no other cell types in the tentacle ectoderm. Battery cells are composed of three distinct regions: the cell body, peripheral attenuated extensions and myonemes. Thick tentacle bands are composed of cell bodies, whereas thin bands are made up of attenuated extensions. Myonemes contribute to both thick and thin regions. It was confirmed that each battery cell has several myonemes, which appear to interdigitate with myonemes of other more proximal and distal battery cells, but not with battery cells of the same BCC ring. Nematocytes have several basal processes. Some processes insert between myonemes and contact the mesoglea; other processes insert into cuplike extensions of myonemes, and are connected to myonemal cups by desmosomal junctions. These observations are discussed in relation to mechanical and electrical aspects of tentacular contraction and bending.     

Head regeneration and polarity reversal inHydra attenuata can occur in the absence of DNA synthesis

Summary Regeneration in hydra is considered to be morphallactic because it can occur in the absence of cell division. Whether DNA synthesis is required for regeneration or other repatterning events is not known. The question was investigated by blocking DNA synthesis with hydroxyurea and examining several developmental processes. Head regeneration, reversal of regeneration polarity and battery cell differentiation all took place in the absence of DNA synthesis. Hence, morphallactic regulation in hydra is independent of both DNA synthesis and mitosis.     

Effects of N-Arachidonoyl Dopamine,Haloperidol, and Their Mixtures on Regeneration of Freshwater Hydra attenuata

N-Arachidonoyl dopamine and haloperidol, both separately and in different combinations, inhibit regeneration of the gastral and basal regions of hydra. In addition, both substances induce stable anomalies of morphogenesis in the form of outgrowths and additional tentacles in gastral regenerates. In the presence of both substances at different combinations, anomalies either do not appear altogether, or exist for a short time, thus suggesting the normalization pf morphogenesis. Possible mechanisms underlying the effects of these substances are discussed.     

Localization of dopamine in the freshwater hydrozoanHydra attenuata

Summary High performance liquid chromatography (HPLC), with electrochemical detection, is an analytical method sensitive enough to permit quantification of dopamine, dihydroxyphenylalanine (DOPA) and 5-S-cysteinyl DOPA in single or hemisected specimens ofHydra attenuata. Dopamine and 5-S-cysteinylDOPA appear to be the quantitatively predominant catechol compounds inH. attenuata, whereas DOPA is present in minor amounts. The presence of DOPA and 5-S-cysteinylDOPA, and the quantitative correlation between dopamine and these compounds in many specimens, suggests that dopamine inH. attenuata, as in higher animals, is formed through decarboxylation of DOPA. Contrary to the dopaminergic nerves in higher animals, DOPA inHydra seems to be oxidized and 5-S-cysteinyl DOPA is formed as a by-product. The oxidation of DOPA indicates that the hydroxylation of tyrosine into DOPA in the tissues ofH. attenuata is mediated by a tyrosinase rather than a tyrosine hydroxylase. Immunocytochemical methods demonstrate a highly variable distribution of dopamine in the tissues of different specimens ofH. attenuata. Dopamine immunoreactivity is confined to ectodermal tissue and can be found in several different cell types including nerve cells, battery cells, nematocytes, epithelial cells and interstitial undifferentiated cells. The large amounts of dopamine found in some specimens ofH. attenuata indicate some biological function, although its sporadic occurrence in neurites makes it less plausible as a generally utilized neurotransmitter in this animal.     

Nucleotide sequence of an actin-encoding gene from Hydra attenuata: structural characteristics and evolutionary implications

We have determined the complete nucleotide sequence of an actin-encoding gene from Hydra attenuata as well as partial sequences of cDNA clones from two additional actin-encoding genes. The gene from the genomic clone contains a single intron, and has promoter and polyadenylation signals similar to those found in other species. The hydra genome has a very A + T-rich base composition (71%). This is reflected in the codon usage of the actin-encoding genes, which is strongly biased towards codons having A or T in the third position. The hydra actin-encoding gene family consists of three or more transcribed genes, two of which are very closely related to each other and probably arose by a recent gene duplication. Hydra actin, like other invertebrate actins, is more similar to the non-muscle isotypes of vertebrates than to the vertebrate muscle actins. Hydra actin is more similar to animal actins than to those of plants or fungi, which is consistent with the view that all metazoans arose from a single protist ancestor.