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At least 1,261 records · Page 70Linked to original sources
PubMed1998

Quantitative analysis of a directed behavior in the medicinal leech: implications for organizing motor output.

The local bend is a directed behavior produced by the leech, Hirudo medicinalis, in response to a light touch. Contraction of longitudinal muscles near the touched location results in a bend directed away from the stimulus. We quantify the relationship between the location of touch around the body perimeter and the behavioral output by using video analysis, muscle tension measurements, and electromyography. On average, the direction of the behavioral output differed from the touch location by <8% of the total body perimeter. We discuss our results in the context of two contrasting behavioral strategies: a Continuous strategy, in which the local bend is directed exactly opposite to stimulus location, and a Categorical strategy, in which there are four distinct bend directions, each elicited by stimuli given in a single quadrant of the body perimeter. To distinguish between these strategies, we delivered two competing stimuli simultaneously. The resulting behavioral output is best described by an average of the effects of each stimulus given alone and thus provides support for the Continuous strategy. We also use a simple model, based on anatomical and physiological data, to predict the responses of the known motor neurons to different stimulus locations. The model shows that the activation of two of the motor neurons (D and V) is inconsistent with a Categorical strategy. However, these neurons are known to be active during the local bend behavior. This result, along with our experimental observations, suggests that the local bend network uses a Continuous strategy to encode stimulus location and produce directed behavioral output.

Animals↗
PubMed1998

Target tissues affect cellular properties of cocultured leech Retzius neurons.

The development of many neurons, including the Retzius (Rz) neurons of the medicinal leech, is shaped in part by interactions with other cells in the environment. To explore the nature of the interaction between growing Rz processes and potential target tissues, adult Rz neurons were cultured directly in contact with some of the tissues that normally serve as their targets in vivo. The morphology of the regenerated processes of these neurons varied depending upon the identity of the target tissues, but other cellular properties remained unchanged. In particular, although during normal development contact with peripheral targets determines the sign of Rz neurons' response to acetylcholine (ACh) applied to the soma, these cultured neurons maintained their original response to ACh even after as long as 2 weeks in culture on novel targets. Hence, some features of cultured adult Rz neurons varied depending upon the conditions, whereas other features remained fixed.

Acetylcholine↗
PubMed1998

Formation of polar cytoplasmic domains (teloplasms) in the leech egg is a three-step segregation process.

Segregation and proliferation of mitochondria, leading to formation of the teloplasms (pole plasms), were studied in eggs of the leech T. rude by immunocytochemistry, fluorescent time lapse video imaging, confocal and electron microscopy. The translocation of mitochondria was analyzed after loading the egg with either Rhodamine 123 or a Mitotracker. Mitochondrial proliferation was assessed after pulse labeling with BrdU. The involvement of the cytoskeleton in the segregation process was determined by drug action. The teloplasms form during the first interphase as consequence of a 3-step sequential process of mitochondrial redistribution throughout the egg cytoplasm. The first step is a microtubule dependent process of ectoplasm thickening due to centrifugal mitochondrial transportation from the neighboring endoplasm. During the second step mitochondria move in the plane of the ectoplasm to become concentrated at the wall of rings (polar rings) and bands of contraction. This process appears to mostly depend on actin. The furrowing pattern of the egg during this step can be modified by cold treatment and seems to be determined during oogenesis. During the third step the ectoplasm flows to either of the poles in conjunction with bipolar displacement of the polar rings and shortening of the contraction bands. This step depends on both microtubules and microfilaments. Mitochondria of first interphase eggs have three special features: (1) they move in clusters, (2) their movement depends on both microtubules and microfilaments and (3) they proliferate continuously. During the first interphase the polarized meiotic egg becomes a bipolar cell.

Actins↗
PubMed1998

Expression of actin mRNA in embryos of the leech Helobdella triserialis.

A cDNA clone (Htr-actin) containing a 1.48 kb insert corresponding to actin was isolated from a stage 10-11 Helobdella triserialis cDNA library by cross hybidization to a Drosophila melanogaster actin clone. The cDNA is equivalent in size to the adult actin mRNA detected by Northern analysis. Genomic Southern blot analysis revealed that Htr-actin is a single copy gene and is one member of a family of actins in H. triserialis. Protein sequence comparisons revealed that Htr-actin was most similar to actin-1 of the earthworm and like other invertebrate actins, Htr-actin was found to be more similar to mammalian cytoplasmic actins than to mammalian muscle-specific actins. In situ hybridization revealed that at early stages the actin mRNA was distributed primarily in the yolk-deficient cytoplasm of all cells. At later stages, elevated levels were detected over the germinal bands and plate. Following muscle formation, a subset of the actin mRNA expression pattern resembled the immunostaining pattern obtained with the muscle specific antibody, Lan3-14. The Htr-actin transcript was undetectable in the segmental nervous system. These studies lay the groundwork for any future ectopic gene expression studies using the Htr-actin promoter in the leech embryo.

Actins↗
PubMed1998

[Internal hirudiniasis: vaginal bleeding resulting from leech bite].

A postmenopausic woman referred continuous bleeding during seven days, which suddenly started after she was swimming in the river. She didn't have a general symptoms. A gynecological examination showed that the cause of the bleeding was a leech bite on the vaginal wall. The worm was gave away by means of surgical forceps without complications. We described the clinical and pathological features of this unusual parasite attack.

Animals↗
PubMed1998

[The leech therapy of infectious myocarditis].

The novelty of the work consists in application of hirudotherapy (leeches) to treat infectious myocarditis, which therapy has been embarked on for the first time in the treatment of this medical condition. Coagulation methods used by the author are modern, the methodical approach is adequate. The role of hirudotherapy versus heparin therapy is shown.

Adolescent↗
PubMed1998

Staging of middle and late embryonic development in the medicinal leech, Hirudo medicinalis.

We present a description of the last half of embryonic development in the European medicinal leech, Hirudo medicinalis, based entirely on externally visible morphological features, and establish reliably observable stages during that development. Embryogenesis, from the time fertilized eggs are deposited in an eggcase (called a cocoon) to the emergence of juveniles from the cocoon, takes approximately 4 weeks at room temperature. The stages described in this paper extend from the completion of segmentation to the appearance of the final bands of pigmentation. Developmental stages are expressed as percentages of total embryonic developmental time. This staging table was constructed for embryos kept at 20 degrees C. In addition, the development of animals kept at 17 degrees C or at 24 degrees C was compared with those held at 20 degrees C. Development proceeds more quickly at higher temperatures. Because development in embryos held at higher or lower temperatures was linearly related to the stages determined for embryos held at 20 degrees C, the rate of development at any intermediate temperature can be predicted from the staging table at 20 degrees C by simple multiplication.

Animals↗
PubMed1998

[Tetrodotoxin effect on spontaneous and evoked impulse activity of leech Retzius neurons].

Investigation into the Retzius neuron response to synaptic low-, medium-, and high-frequency stimulation under the block of the TTX-sensitive sodium inflow channels, was performed in leeches. The TTX block affected the spontaneous unit activity and not the evoked activity, it altered the responses to low-frequency stimulation (sensitisation) and eliminated the responses to high-frequency activation (habituation). The findings suggest the fundamental plastic properties of the Retzius neurons to depend on functional characteristics of the TTX-sensitive sodium channels.

Action Potentials↗
PubMed2005

5-Hydroxytryptamine activates a barium-sensitive, cAMP-mediated potassium conductance in the leech giant glial cell.

5-Hydroxytryptamine (5-HT), a neurotransmitter and neuromodulator in the central nervous system of the leech Hirudo medicinalis hyperpolarizes the giant glial cell in the neuropil of segmental ganglia at micromolar concentrations. The 5-HT-evoked glial response (EC(50) approximately 2.5 microM) is mediated by a non-desensitizing, G-protein-coupled receptor and due to activation of a Ca(2+)-independent K(+) conductance. The adenylyl cyclase inhibitor SQ22,536 blocks the response to 5-HT; in the presence of 1 mM db-cAMP, but not of 1 mM db-cGMP, the glial response is suppressed. The 5-HT-evoked response is reduced by Ba(2+) with half-maximal inhibition at 50 microM Ba(2+). The results suggest that release of 5-HT from serotonergic neurons, or the maintenance of micromolar levels of extracellular 5-HT in the ganglion, may help to set the glial membrane potential close to the K(+) equilibrium potential.

Animals↗
PubMed2006

5-Hydroxytryptamine-mediated increase in glutamate uptake by the leech giant glial cell.

The clearance of synaptically released glutamate is one of the pivotal functions of glial cells. We have studied the role of 5-hydroxytryptamine (5-HT, 30 microM), a neurotransmitter and neurohormone in the leech central nervous system with a versatile action spectrum, on the efficacy of glial glutamate uptake. The activity of the glutamate uptake carrier in the giant glial cell in isolated ganglia of Hirudo medicinalis was monitored by measuring the membrane current and the change in the intracellular Na(+) concentration (Na(+) (i)) as induced by the glutamate carrier substrate D-aspartate (D-asp, 1 mM). 5-HT increased the D-asp-induced current (EC(50) at 5 microM) and rise in Na(+) (i), an effect which was mimicked by the membrane-permeable cyclic nucleotide analogue dibutyryl-cyclic AMP (db-cAMP). The adenylyl cyclase inhibitor SQ 22,536 and the protein kinase A antagonist Rp-cAMP inhibited the effect of 5-HT. Blocking the G protein in the giant glial cell by injecting GDP-beta-S suppressed the effect of 5-HT, but not the effect of db-cAMP, on the D-asp-induced current. Our results suggest that 5-HT enhances the glial uptake of glutamate via cAMP- and PKA-mediated pathway.

Adenylyl Cyclase Inhibitors↗
PubMed1999

Wound healing after silver nitrate burns in the leech hirudo medicinalis

The healing process after inflicting silver nitrate burns, a type of injury in which a heavy loss of body wall tissues occurs, was studied in Hirudo medicinalis. Silver solution penetrates the body wall and silver precipitates in the tissues during infiltration. Vasocentral cells, do not penetrate the impregnated zone but accumulate just below it to form a pseudoblastema. The impregnated zone splits off entirely, leaving an area covered by pseudoblastema cells on the leech surface, which is progressively reepithelialized. There is no regeneration of the lost body region, and although some connective tissue matrix may be produced by pseudoblastema cells, there is no significant regeneration of the connective tissue and nor the muscular fibers of the body wall, resting an evident depression in the damaged zone. The amount of affected tissue, and possibly the inability of vasocentral cells to reach the infiltrated tissues are determinant factors in the development of the splitting process. Copyright 1999 Academic Press.

Journal Article↗
PubMed2005

Development of swimming in the medicinal leech, the gradual acquisition of a behavior.

Observing the development of behavior provides an assay for the developmental state of an embryo's nervous system. We have previously described the development of behaviors that were largely confined to one or a few segments. We now extend the work to a kinematic analysis of the development of swimming, a behavior that requires coordination of the entire body. When leech embryos first begin to swim they make little forward progress, but within several days they swim as effectively as adults. This increase in efficacy depends on changes in body shape and on improved intersegmental coordination of the swim central pattern generator. These kinematic details suggest how the swim central pattern generating circuit is assembled during embryogenesis.

Age Factors↗
PubMed2006

Postactivation inhibition of spontaneously active neurosecretory neurons in the medicinal leech.

Spontaneously active neurosecretory neurons in vertebrate and invertebrate nervous systems share similarities in firing frequencies, spike shapes, inhibition by the transmitters they themselves release and postactivation inhibition, an intensity-dependent period of suppressed spontaneous generation of action potentials following phases of high-frequency activity. High-frequency activation of spontaneously active serotonin-containing Retzius cells in isolated ganglia of the leech Hirudo medicinalis induced prolonged membrane hyperpolarisations causing periods of postactivation inhibition of up to 33 s. The duration of the inhibitory periods was directly related to both the number and rate of spikes during activation and was inversely proportional to a cell's spontaneous firing frequency. The periods of postactivation inhibition remained unaffected by both serotonin depletion through repeated injections of 5,7-dihydroxytryptamine and suppressing the afterhyperpolarisation following each action potential with tetraethylammonium (TEA), iberiotoxin or charybdotoxin, suggesting that neither autoinhibition by synaptic release of serotonin nor calcium-activated potassium channels contribute to the underlying mechanism. In contrast, the postactivation inhibitory period was significantly affected both by differential electrical stimulation of the same Retzius cells via microelectrodes filled with molar concentrations of either Na(+)-acetate or K(+)-acetate, and by partial inhibition of Na(+)/K(+)-ATPase with ouabain. Thus, postactivation inhibition in Retzius cells results from prolonged hyperpolarising activity of Na(+)/K(+)-ATPase stimulated by the accumulation of cytosolic Na(+ )during phases of high-frequency spike activity.

Action Potentials↗
PubMed2005

Ionic mechanism of ouabain-induced swelling of leech Retzius neurons.

By using electrophysiological and microfluorimetric methods, we found that leech Retzius neurons swell after inhibition of the Na(+)-K(+) pump by the cardiac glycoside ouabain. To explore the mechanism of this swelling, we measured the effect of ouabain on [Na(+)](i), [K(+)](i), and [Cl(-)](i), as well as on the membrane potential, by applying triple-barrelled ion-sensitive microelectrodes. As shown previously, ouabain induced a marked [Na(+)](i) increase, a [K(+)](i) decrease, and a membrane depolarization, and it also evoked an increase in [Cl(-)](i). The analysis of the data revealed a net uptake of NaCl, which quantitatively explained the ouabain-induced cell swelling. In the absence of extracellular Na(+) or Cl(-), NaCl uptake was excluded, and the cell volume remained unaffected. Likewise, NaCl uptake and, hence, cell swelling did not occur when the Na(+)-K(+) pump was inhibited by omitting bath K(+). Also, in K(+)-free solution, [Na(+)](i) increased and [K(+)](i) dropped, but [Cl(-)](i) slightly decreased, and after an initial, small membrane depolarization, the cells hyperpolarized for a prolonged period. It is concluded that the ouabain-induced NaCl uptake is caused by the depolarization of the plasma membrane, which augments the inwardly directed electrochemical Cl(-) gradient.

Animals↗
PubMed2006

Activity-dependent suppression of spontaneous spike generation in the Retzius neurons of the leech Hirudo medicinalis L.

We report on factors affecting the spontaneous firing pattern of the identified serotonin-containing Retzius neurons of the medicinal leech. Increased firing activity induced by intracellular current injection is followed by a 'post-stimulus-depression' (PSD) without spiking for up to 23 s. PSD duration depends both on the duration and the amplitude of the injected current and correlates inversely with the spontaneous spiking activity. In contrast to serotonin-containing neurons in mammals, serotonin release from the Retzius cells presumably does not mediate the observed spike suppression in a self-inhibitory manner since robust PSD persists after synaptic isolation. Moreover, single additional spikes elicited at specific delays after spontaneously occurring action potentials are sufficient to significantly alter the firing pattern. Since sub-threshold current injections do not affect the ongoing spiking pattern and PSD persists in synaptically isolated preparations our data suggest that PSD reflects an endogenous and 'spike-dependent' mechanism controlling the spiking activity of Retzius cells in a use-dependent way.

Animals↗
PubMed2004

Sensitization and dishabituation of swim induction in the leech Hirudo medicinalis: role of serotonin and cyclic AMP.

In this paper the role of serotonin (5HT) and cyclic AMP (cAMP) in sensitization and dishabituation of swim induction (SI) has been investigated in the leech Hirudo medicinalis. Electrical stimulation of the body wall evokes swimming activity with a constant latency. In animals with a disconnection between head ganglion and segmental ganglia, repetitive stimulation induces habituation of swimming whereas brushing on the dorsal skin provokes sensitization of a naïve response or dishabituation of a previously habituated response. Our findings indicate that 5HT is the neurotransmitter underlying both sensitization and dishabituation of SI. Injection of the 5HT receptor blocking agent methysergide impaires the onset of sensitization and dishabituation induced by brushing. Moreover, injection of 5HT mimics these forms of nonassociative learning, whereas injection of dopamine does not. Finally, the effects of 5HT are mediated by cAMP: (1) after injections of specific adenylate cyclase inhibitors such as MDL 12.330A or SQ22536, brushing becomes ineffective in facilitating the SI in either non-habituated or habituated animals. (2) 8Br-cAMP application mimics both sensitization and dishabituation of SI.

Animals↗
PubMed2005

Study of a major intermediate in the oxidative folding of leech carboxypeptidase inhibitor: contribution of the fourth disulfide bond.

The oxidative folding pathway of leech carboxypeptidase inhibitor (LCI; four disulfide bonds) proceeds through the formation of two major intermediates (III-A and III-B) that contain three native disulfide bonds and act as strong kinetic traps in the folding process. The III-B intermediate lacks the Cys19-Cys43 disulfide bond that links the beta-sheet core with the alpha-helix in wild-type LCI. Here, an analog of this intermediate was constructed by replacing Cys19 and Cys43 with alanine residues. Its oxidative folding follows a rapid sequential flow through one, two, and three disulfide species to reach the native form; the low accumulation of two disulfide intermediates and three disulfide (scrambled) isomers accounts for a highly efficient reaction. The three-dimensional structure of this analog, alone and in complex with carboxypeptidase A (CPA), was determined by X-ray crystallography at 2.2A resolution. Its overall structure is very similar to that of wild-type LCI, although the residues in the region adjacent to the mutation sites show an increased flexibility, which is strongly reduced upon binding to CPA. The structure of the complex also demonstrates that the analog and the wild-type LCI bind to the enzyme in the same manner, as expected by their inhibitory capabilities, which were similar for all enzymes tested. Equilibrium unfolding experiments showed that this mutant is destabilized by approximately 1.5 kcal mol(-1) (40%) relative to the wild-type protein. Together, the data indicate that the fourth disulfide bond provides LCI with both high stability and structural specificity.

Animals↗
PubMed2005

NMR structural characterization and computational predictions of the major intermediate in oxidative folding of leech carboxypeptidase inhibitor.

The III-A intermediate constitutes the major rate-determining step in the oxidative folding of leech carboxypeptidase inhibitor (LCI). In this work, III-A has been directly purified from the folding reaction and structurally characterized by NMR spectroscopy. This species, containing three native disulfides, displays a highly native-like structure; however, it lacks some secondary structure elements, making it more flexible than native LCI. III-A represents a structurally determined example of a disulfide-insecure intermediate; direct oxidation of this species to the fully native protein seems to be restricted by the burial of its two free cysteine residues inside a native-like structure. We also show that theoretical approaches based on topological constraints predict with good accuracy the presence of this folding intermediate. Overall, the derived results suggest that, as it occurs with non-disulfide bonded proteins, native-like interactions between segments of secondary structure rather than the crosslinking of disulfide bonds direct the folding of LCI.

Animals↗