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PubMed1983

Inhibition of lung tumor colonization by leech salivary gland extracts from Haementeria ghilianii.

Salivary gland extract from the South American leech Haementeria ghilianii, administered i.v. on the same day as the i.v. inoculation of T241 sarcoma cells, completely suppresses colonization of the mediastinal lymph nodes and markedly reduces the number and size of lung tumor colonies produced by this tumor. Additional studies indicate that the extract contains various types of proteinase inhibitors and has the capacity to inhibit clotting and platelet aggregation by tumor material and collagen. Although not yet proved by direct evidence, these activities may be involved in the inhibitory effect of lung tumor colonization by the leech extract.

Animals↗
PubMed1982

Segmental specialization of a leech swim-initiating interneuron, cell 2051.

The physiological and anatomical properties of an unpaired intersegmental interneuron designated cell 205 are described. Cell 205 is unusual among leech neurons in that it combines a variety of functional properties in a single cell. Constant current depolarization of cell 205 initiates and maintains swimming behavior in semi-intact leeches or the swim motor pattern in brainless, isolated nerve cords. During swim episodes elicited by other stimuli, cell 205 is rhythmically active. Current pulses passed into cell 205 during swimming reset the pattern, indicting that it has access to, or may be a member of, the swim central pattern generator (CPG). Cell 205 is the first interneuron in this system to exhibit both swim-initiating and CPG functions. Individual touch, pressure, and nociceptive primary mechanoreceptor neurons polysynaptically excite cell 205, which, in addition, is coupled electrically to the multimodal S interneuron. These inputs may contribute to the initiation and/or modulation of swimming in response to sensory stimuli. Cell 205 shares some common synaptic inputs and outputs with the only other known swim-initiating interneuron, cell 204, but the two cells differ fundamentally in that cell 204 exerts only a tonic effect on the CPG. No synaptic interactions were found between cells 204 and 205, but their excitatory effects on swimming summate. Unlike other swim neurons which are segmentally repeated, cell 205 generally is present only in segment 9, and numerous lines of evidence suggest that it is, in fact, a segmentally differentiated homolog of cell 204.

Animals↗
PubMed1994

Distribution of carbohydrate epitopes among disjoint subsets of leech sensory afferent neurons.

Carbohydrate recognition plays an important role in the development of normal projections of sensory afferent neurons in the leech CNS. Four different carbohydrate epitopes are expressed by sensory afferents on their 130 kDa surface proteins: all sensory afferents share a common carbohydrate epitope (CE0) that helps them to enter and project diffusely across the synaptic neuropil; a restricted expression of three other carbohydrate epitopes (CE1, CE2, and CE3) serves to distinguish three subsets of sensory afferents. We examined the subsets of sensory afferents defined by their subset carbohydrate epitopes in the leech lip, skin, gut, and CNS. We established that the CE1, CE2, and CE3 subset epitopes define disjoint subsets of neurons by double labeling sensory afferents with monoclonal antibodies for different pairs of subset epitopes. We found that CE2 and CE3 afferents populate the lip and skin, but not the gut, and that these two subsets of sensory afferents have convergent projection patterns in the CNS. We found that CE1 afferents populate the gut and skin, but not lips; furthermore, their CNS projections diverge from those of CE2 and CE3 afferents. Our data fit the hypothesis that these carbohydrate epitopes are related to sensory modality of afferent subsets.

Animals↗
PubMed1995

Two kinetically distinct 5-hydroxytryptamine-activated Cl- conductances at Retzius P-cell synapses of the medicinal leech.

The properties of serotonin (5-HT)-activated Cl- receptor/ion channel complexes in neurons of the CNS of the medicinal leech were analyzed. These channels mediate the postsynaptic response at the serotoninergic Retzius P-cell as well as Retzius-Retzius cell synapses. 5-HT-induced Cl- currents were activated by fast superfusion of transmitter on cells dissociated from embryonic leeches including histochemically identified Retzius cells. Whole-cell currents elicited by partial superfusion of the membrane and summated single-channel currents of outside-out patches showed times to peak of 18 +/- 4 msec and 10 +/- 5 msec, respectively, and desensitized with time constants of 28 +/- 3 msec and 20 +/- 11 msec. Persistence of single-channel openings in the outside-out configuration as well as lack of effect of dialysis of the whole cell with AMP-PNP or GDP-beta-S indicated that 5-HT directly gates the Cl- channels without involving second messenger cascades. In outside-out patches, two single-channel conductances of 13 pS and 32 pS were identified. While the 13 pS conductance desensitized, the 32 pS conductance activated within several tens of msec and showed no desensitization. We postulate that two subtypes of channels are coactivated by 5-HT and that the activation of the fast desensitizing channel could be responsible for the fast decaying component of the post-synaptic response. The slow conductance explains the second slower decay time constant of the postsynaptic response and could account for the tonic component sometimes observed at Retzius P-cell synapses.

Animals↗
PubMed1994

Segmental specificity of neuronal recognition during synapse formation between identified leech neurons.

An early event in the formation of inhibitory synapses between the serotonergic Retzius (R) neuron and the pressure-sensitive (P) neuron of the leech is the elimination of an extrasynaptic, depolarizing response to 5-HT in the P cell from sites of contact. This effect is induced specifically by contact with the R neuron but not with other identified leech neurons, including other serotonergic neurons that do not form chemical synapses with the P cell. In the reproductive (fifth and sixth) segmental ganglia, the R cells (R(5,6)) exhibit functional and morphological differences with the standard R cells (R(x)) in the other 19 segmental ganglia. In the study presented here, we tested the specificity of P cell recognition of R cells by examining whether there were differences between the R(x) and R(5,6) cells with respect to synapse formation with the P cell. R(5,6) neurons did not innervate P cells in the fifth and sixth ganglia (P(5,6)) in vivo or in vitro, nor did they form synapses with P cells from standard ganglia (P(x)) in vitro except on rare occasions, after long periods in culture. In contrast to the effects of R(5,6) neurons, R(x) neurons readily innervated both P(x) and P(5,6) cells in vitro, suggesting that the lack of an R(5,6)-P(5,6) synapse reflects differences in the presynaptic rather than the postsynaptic cell. In contrast to the R(x) cell, contact with the R(5,6) neurons had no significant effect on the responses of the P cell to 5-HT.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

The development of a hemogregarine of Lycodes raridens from Alaska in its definitive leech host.

Piscicolid leeches from the Bering Sea that had been removed from Lycodes raridens in 1977 were examined histologically. Developmental stages of a haemogregarine blood parasite of fish were found in the intestinal ceca and in salivary tissues. The developmental sequence, including gametogenesis and sporogony followed by intestinal merogonic development and anteriad migration, was the same as is known for Haemogregarina (sensu lato) myoxocephali of Atlantic sculpins. Merozoites with perinuclear crystalloid inclusions were found in the dorsal blood vessel, interstitially between the blood vessel and salivary cells and within both the somata and ductules of salivary cells. This confirms the importance of rhynchobdellid leech circulatory anatomy in facilitating the transmission of apicomplexan parasites. The lack of distinguishing characteristics precludes assigning a specific name to this parasite and may be indicative of frustrations that will be encountered in future revisions of the systematics of the genus Haemogregarina.

Animals↗
PubMedDate not supplied

[Unavoidable epistaxis in the nasal infestation of leeches].

An adult man is presented with a case of epistaxis due to leech (Limnatis linotica) intranasal parasitation. The geographical area where infection was performed is in the province of Teruel, northwest Spain. The only presented symptom was an incoercible nasal haemorrhage with low flow which didn't cease by anterior tamponade. 7 days after the process began he was heeded at the Emergency Service, Alcañiz Hospital, where the exam of left nasal fossa showed the presence of a olive-green long mass, very mobile. The treatment consisted on the leech extraction with a dissection forceps. Haemorrhage stopped immediately after the mild intervention and, reexamination the nasal fossa 15 days after, it was no wound or infectious complication observed. The patient was infested when drinking water from a fountain placed in rural area.

Aged↗
PubMed1993

Relying on past science: nursing implications of medicinal leech therapy.

The resurgence of leech therapy is due to a need to re-establish venous circulation after micro-surgical procedures such as the reattachment of amputated body parts. Using a medicinal leech, Hirudo medicinalis, this painless procedure can drain up to 20 ml of blood at feeding and an additional amount via slow but continuous flow after feeding. Nurses play a key role before, during, and after the therapy in educating and emotionally preparing patients and families. Guidelines for nursing actions during the therapy as well as implications for nursing research are discussed.

Animals↗
PubMed1993

Persistence of leech repellents on cloth.

Trials on persistence of repellent properties of N, N-diethyl phenyl acetamide (DEPA), N,N-diethyl-m-toluamide (DEET), 3acetyl2(2-6-dimethyl-5-heptenyl)oxazolidine(Citronyl) , dimethyl phthalate (DMP) and N-benzoyl piperidine (NBP) on cloth were conducted against land leeches in evergreen rain and deciduous forests of Assam. Results obtained were compared with volatile oil of Zanthoxylum armatum DC. syn. Z. alatum Roxb (Timur) to evaluate its efficacy as leech repellent. DEPA and DEET were found to be the best. Timur oil was at par with Citronyl and exhibited better results than DMP and NBP.

Acetamides↗
PubMed1993

Cytopathological changes induced by Haemogregarina myoxocephali in its fish host and leech vector.

The cytopathological effects of infection with Haemogregarina myoxocephali were investigated by light and electron microscopy in its fish host, Myoxocephalus octodecemspinosus, and in its leech vector, Malmiana scorpii. Gamont-infected erythrocytes in fish appeared vacuolated and exhibited shrinkage and altered shape consistent with loss of cytoskeletal integrity. Infected intestinal epithelial cells in leeches showed changes suggestive of recruitment of host metabolic functions by the developing parasite, including hypertrophied mitochondria and accumulations of endoplasmic reticulum and secretory vesicles next to the parasitophorous vacuole.

Animals↗
PubMed1995

[Investigation and experimental study on the standards and quality of leech pieces].

It has been confirmed by investigation that Whitmania pigra is the main medicinal breed of leech, and filiform pieces are the best for extraction. The quality of leech processed by 3 different methods has been studied qualitatively and quantitatively in terms of amounts of extraction, TLC, electrophoresis of protein and anti-thrombin action.

Animals↗
PubMed1997

The neuronal basis of the behavioral choice between swimming and shortening in the leech: control is not selectively exercised at higher circuit levels.

Swimming and the whole-body shortening reflex are two incompatible behaviors performed by the medicinal leech Hirudo medicinalis. We set out to examine the neuronal basis of the choice between these behaviors, taking advantage of the fact that the neuronal circuit underlying swimming is relatively well understood. The leech swim circuit is organized hierarchically and contains three interneuronal levels, including two upper levels of "command-like" neurons. We tested the responses of the swim circuit neurons to stimuli that produced shortening, using reduced preparations in which neurophysiological recording could be performed while behaviors were elicited. We found that the majority of the swim circuit neurons, including most of the command-like cells and all of the cells at the highest hierarchical level of the circuit, were excited by stimuli that produced shortening as well as by stimuli that produced swimming. Only a subset of neurons, at levels below the top, were inhibited during shortening; these included one of the command-like cells and an oscillator cell (an interneuron that is part of the central pattern generator for swimming). These results imply that the control of the choice between swimming and shortening is not exercised selectively at the higher levels of the swim circuit.

Action Potentials↗
PubMed1997

Intracellular acidification of the leech giant glial cell evoked by glutamate and aspartate.

Glutamate is an excitatory receptor agonist in both neurones and glial cells, and, in addition, glutamate is also a substrate for glutamate transporter in glial cells. We have measured intracellular and extracellular pH changes induced by bath application of glutamate, its receptor agonist kainate, and its transporter agonist aspartate, in the giant neuropile glial cell in the central nervous system of the leech Hirudo medicinalis, using double-barrelled pH-sensitive microelectrodes. The giant glial cells responded to glutamate and aspartate (100-500 microM), and kainate (5-20 microM) with a membrane depolarization or an inward current and with a distinct intracellular acidification. Glutamate and aspartate (both 500 microM) evoked a decrease in intracellular pH (pHi) by 0.187 +/- 0.081 (n = 88) and 0.198 +/- 0.067 (n = 86) pH units, respectively. With a resting pHi of 7.1 or 80 nM H+, these acidifications correspond to a mean increase of the intracellular H+ activity by 42 nM and 45 nM. Kainate caused a decrease of pHi by 0.1-0.35 pH units (n = 15). The glutamate/aspartate-induced decrease in pHi was not significantly affected by the glutamate receptor blockers kynurenic acid (1 mM) and 6-cyano-7-dinitroquinoxaline-2,3-dione (CNQX, 50-100 microM), which greatly reduced the kainate-induced change in pHi. Extracellular alkalinizations produced by glutamate and aspartate were not affected by CNQX. Reduction of the external Na+ concentration gradually decreased the intracellular pH change induced by glutamate/aspartate, indicating half maximal activation of the acidifying process at 5-10 mM external Na+ concentration. When all external Na+ was replaced by NMDG+, the pHi responses were completely suppressed (glutamate) or reduced to 10% (aspartate). When Na+ was replaced by Li+, the glutamate- and aspartate-evoked pHi responses were reduced to 18% and 14%, respectively. Removal of external Ca2+ reduced the glutamate- and aspartate-induced pHi responses to 93 and 72%, respectively. The glutamate/aspartate-induced intracellular acidifications were not affected by the putative glutamate uptake inhibitor amino-adipidic acid (1 mM). DL-aspartate-beta-hydroxamate (1 mM), and dihydrokainate (2 mM), which caused some pHi decrease on its own, reduced the glutamate/aspartate-induced pHi responses by 40 and 69%, respectively. The putative uptake inhibitor DL-threo-beta-hydroxyaspartate (THA, 1 mM) induced a prominent intracellular acidification (0.36 +/- 0.05 pH units, n = 9), and the pHi change evoked by glutamate or aspartate in the presence of THA was reduced to less than 10%. The results indicate that glutamate, aspartate, and kainate produce substantial intracellular acidifications, which are mediated by at least two independent mechanisms: 1) via activation of non-NMDA glutamate receptors and 2) via uptake of the excitatory amino acids into the leech glial cell.

2-Aminoadipic Acid↗
PubMed1997

The establishment of peripheral sensory arbors in the leech: in vivo time-lapse studies reveal a highly dynamic process.

Pressure-sensitive (P) neurons located in the leech CNS form elaborate terminal arbors in the body wall of the animal during mid-embryogenesis. In the experiments discussed here, arbor development in the target region was studied in intact, unanesthetized leech embryos using time-lapse video microscopy of individual, fluorescently stained P neurons. Analysis of time-lapse recordings made over a period of several days revealed that arbor formation is a very dynamic process. At any particular time, most high-order terminal branches were either extending or retracting, in approximately equal numbers and at very similar rates. Many branches underwent several rounds of extension and retraction every hour. Net arbor growth occurred at a much lower rate than the extension and retraction rates of individual branches. Process retraction sometimes resulted in an apparent change in the topological order of processes. Significantly, the initiation of new branches was restricted to a few locations along the parent process, which were termed "hot spots." Moreover, the capacity to generate high-order branches correlated with parent process stability. The target region of the growing P cell arbor in the body wall was subsequently examined using confocal microscopy in fixed preparations. The arbor expanded between the longitudinal and circular muscle layers, a region occupied by small unidentified cells. Simultaneous imaging of the dye-labeled terminal arbor and the surrounding tissue at two different wavelengths suggested that the high-order processes were navigating around these cells, which sometimes forced the growing processes to assume a bent form. These observations suggest that the formation of the P cell arbor can be best described as a "dynamically unstable" process that is constrained by interactions with its environment.

Animals↗
PubMed1998

Development and pathway formation of peripheral neurons during leech embryogenesis.

By labeling the germinal plates of staged leech embryos with monoclonal antibodies to the immunoglobulin superfamily member Tractin, we have documented the distribution and initial development of peripheral neurons in a hirudinid leech. We find, in addition to sensillar and extrasensillar sensory neurons, that there are 21 identifiable peripheral neurons in each hemisegment. These neurons are found in highly stereotyped positions, and all but two of them are associated with the segmental nerves. We show that eight of the peripheral neurons have the characteristic morphology of stretch receptor neurons and that they form a circumferentially distributed grid aligned in such a way that each of five specialized longitudinal muscle fascicles are monitored by at least two stretch receptor cells covering ventral, lateral, and dorsal regions of the body wall. Furthermore, we show that, in contrast to the dorsal posterior nerve, which is pioneered by central projections, the pathways of the three remaining segmental nerves are likely to be pioneered or guided by peripheral neurons.

Animals↗
PubMed1998

Fibrino(geno)lytic properties of purified hementerin, a metalloproteinase from the leech Haementeria depressa.

The fibrino(geno)lytic protein designated hementerin contained in crude extracts of the salivary complex of Haementeria depressa leeches was purified to apparent homogeneity by gel filtration, ion exchange chromatography and preparative SDS-PAGE. It is a single-chain 80 kDa, PhMeSO2F-resistant, calcium-dependent, metalloproteinase, which specifically degrades fibrin(ogen) through a plasminogen-independent pathway. The amino terminal sequence of 8 residues shows 80% similarity with hementin, another fibrino(geno)lytic protein purified from Haementeria ghilianii leeches. However, their activities differ somewhat in terms of kinetics and with regard to the structure of the fibrin(ogen) fragments they may produce. Cleavage by hementerin of fibrinogen Aalpha, gamma and Bbeta chains, in that order, produces 270 kDa to 67 kDa fragments which differ from those produced by plasmin. Hementerin was also able to degrade cross-linked fibrin although at a lower rate as compared to fibrinogen. In conclusion, hementerin is a plasminogen-independent fibrino(geno)lytic metalloproteinase that degrades fibrinogen faster than fibrin, prevents blood coagulation and destroys fibrin clots in vitro.

Animals↗
PubMed1998

Long-lasting depolarization of leech neurons mediated by receptors with a nicotinic binding site.

The serotonergic Retzius neurons of the leech midbody ganglia respond in a complex manner to pressure pulses of acetylcholine (ACh) applied onto their soma with a fast depolarization followed by a slower hyperpolarization and an additional delayed long-lasting depolarization. The delayed depolarization is the subject of the present study. The delayed depolarization could be elicited by long (> 1 s) ACh pressure pulses or by short pulses (10 ms) of carbachol, nicotine and DMPP, but not by muscarinic agonists. It was inhibited by bath application of nicotine (10-100 mumol l-1), strychnine (100 mumol l-1) and atropine (10-100 mumol l-1). Nicotinic antagonists that blocked the fast depolarization and the slow hyperpolarization (100 mumol l-1 mecamylamine and d-tubocurarine) did not affect the delayed depolarization induced by carbachol. Partial replacement of the extracellular Na+ by glucamine caused a decrease in the amplitude of the response and a shift of its reversal potential to more negative values. Carbachol pulses applied to Retzius neurons of the ganglia innervating the reproductive segments elicited delayed depolarizations of much smaller amplitude than the ones recorded in Retzius neurons from standard segments. The delayed depolarization could be elicited by the application of short agonist pulses onto different loci over the surface of the ganglion, at a distance from the soma. Isolated cultured Retzius neurons did not exhibit the delayed depolarization although they readily expressed the earlier phases of the complex cholinergic response. Carbachol pulses applied to the soma of other neurons in the leech ganglion produced a variety of specific responses. The results suggest that the delayed depolarization was produced by the activation of a cationic conductance mediated by receptors with a pharmacological profile similar to that of the alpha 9 nicotinic receptors and was not a byproduct of the early phases of the cholinergic response. The response seemed to be initiated in the extensive neuropilar processes of the Retzius cell, enabling a persistent excitatory signal.

Acetylcholine↗
PubMed1998

Leech antihemostatic proteins share the T-knot scaffold, a disulfide-reinforced structural motif.

The occurrence of similar topologies among unrelated proteins is an emerging theme in structural biology. Here we report that the T-knot scaffold, a disulfide-reinforced structural motif shared by knottins and EGF-like proteins, is also present in leech antihemostatic proteins. Our finding emphasizes the versatile nature of this small structural motif, representing a compact structural unit suitable for the diverse biological functions performed by knottins, EGF-like proteins and leech antihemostatic proteins.

Animals↗