Ventral in position, located at the base of the buccal cavity. Dissection of Nervous System: Fix the head of the speci­men by pinning through the mandibles. In 1989 and 1990 two articles describing the application of patch-clamp technique in the study on the activity of neurosecretory dorsal unpaired median (DUM—Figure 1(c)) neurons from terminal ganglion of Periplaneta americana nerve cord were published [44, 45]. In the presence of the toxin, resting depolarization was observed jointly with a tendency to repetitive discharges. 1. The name “depressant toxins” comes from the flaccid paralysis they cause in fly larvae. Alpha-like toxins do not express selectivity toward insect or mammalian sodium channels. The activity of giant axon can also be recorded in in situ conditions in the whole nerve cord [7]. Nerves arising from sub-oesopha­geal ganglia end in labrum, mandible and both pairs of maxillae. At the presynaptic face of such synapses, muscarinic receptors are present, the role of which is to regulate the acetylcholine releasing by negative feedback [62]. Also learn about:- 1. Biophysics Laboratory, Institute of General and Molecular Biology, N. Copernicus University, 87-100 Torun, Poland, Departamento de Bioquímica e Imunologia, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Belo Horizonte, 31270-901 MG, Brazil. Sodium current was activated at more negative potential than normally; it was prolonged, but holding current development was slower than in the case of depressant toxins [31]. At the same time, its affinity to sodium channel target and its toxicity decreased. The rectum opens through the anus. P. Lazarovici, P. Yanai, M. Pelhate, and E. Zlotkin, “Insect toxic components from the venom of a chactoid scorpion, L. Borchani, P. Mansuelle, M. Stankiewicz et al., “A new scorpion venom toxin paralytic to insects that affects Na, L. Borchani, M. Stankiewicz, C. Kopeyan et al., “Purification, structure and activity of three insect toxins from, R. Benkhalifa, M. Stankiewicz, B. Lapied et al., “Refined electrophysiological analysis suggests that a depressant toxin is a sodium channel opener rather than a blocker,”, R. B. Khalifa, M. Stankiewicz, M. Pelhate et al., “Action of babycurus-toxin 1 from the east african scorpion, M. Stankiewicz, F. Grolleau, B. Lapied, L. Borchani, M. El Ayeb, and M. Pelhate, “Bot it2, a toxin paralytic to insects from the. The two oviducts join posteriorly and form a muscular tube, the vagina. Along with the science progressing, modern techniques in molecular biology were applied and a “new era” in scorpion anti-insect toxins launched. The first observed effect, after the toxin’s application, was a repetitive generation of slightly prolonged action potentials in response to short stimulation—such effect has never been observed in the experiments on isolated giant axon (Figure 4(c)). F. Grolleau, M. Stankiewicz, L. Birinyi-Strachan et al., “Electrophysiological analysis of the neurotoxic action of a funnel-web spider toxin, B. Lapied, M. Stankiewicz, F. Grolleau, H. Rochat, E. Zlotkin, and M. Pelhate, “Biophysical properties of scorpion alpha-toxin-sensitive background sodium channel contributing to the pacemaker activity in insect neurosecretory cells (DUM neurons),”, M. Stankiewicz, F. Grolleau, H. Rochat, E. Zlotkin, M. Pelhate, and B. Lapied, “Insect background sodium channel as a new target for scorpion alpha toxin,”, F. Grolleau, M. Stankiewicz, E. Kielbasiewicz et al., “Indirect activation of neuronal noncapacitative Ca. Classical voltage-dependent sodium channels were also modified in a manner typical for beta toxin. In the experiments on the isolated axon of the cockroach, it has been shown that the replacement of the amino acid in position 58 from Asn to Asp is able to change completely the toxin’s mode of action. Using the iontophoretic application of acetylcholine or carbachol to postsynaptic membrane vicinity permits to discriminate between pre- and post-synaptic action of the tested drugs. The application of alpha toxin (LqhαIT, 10−5 and 10−4 M) never induced plateau action potential in situ; only transformation from regular firing into bursting activity was observed and the bursts were generated from a level of slight depolarization (Figure 5(c)). Evoked responses are generated by electrical stimulation of the cercal nerve or direct mechanical stimulation of the cerci. POKL.04.01.01-00-081/10),” Poland. Expose the roots of the circumoesophageal connectives on the lateral sides of the brain and trace them to the points where they meet the sub-oesophageal ganglia (Fig. Carefully remove all the tra­cheae and fat in the region where the salivary apparatus is lodged. Endocrine System […] 6.1) with your left hand and clip the wings. In the toxin’s presence, single-amplitude openings of the channels were replaced by events with several distinct subconductance current levels. Put clear water in the tray. Dissection of Reproductive System. Paired, located under the 4th and 5th abdominal terga. 6.2). Exalted axonal excitability resulted from the increase in sodium current amplitude at negative membrane potentials and its prolongation observed under voltage pulse (Figure 3(b)). New, recombinant toxins LqhαITr [36], Lqh IT2 [37] and Bj-xtrIT [38], tested on the cockroach isolated giant axon, showed much resemblance in their mode of action to the native counterparts. Carefully lift it with a spatula and place in a watch glass containing water. A narrow tube, divisible into 3 zones – ileum, colon and rectum. The paired testes are situated just beneath the 4th and 5th abdomi­nal terga. Confronting the results obtained reveals that a toxin may affect the activity of the same nervous structure with diverse effects, depending on the experimental conditions and this conclusion should be taken into account prior to any definite statement concerning the mode of action of any toxin. From hypo-cerebral gan­glia nerves go to corpora cardiaca or oesphageal ganglia. The influence of axonal microenvironment created by glial cells and other axons is a very likely factor in this matter. Using a virtual dissection allows us to take a closer look at body systems using technology such as … It is the anterior most and largest part […] Cockroach model has been largely used for the description of their mode of action. Such studies provided an extremely valuable conclusion: the activity of properly prepared recombinant toxins is the same as that of the native ones. 6.3). Recurrent nerve connects frontal and hypo-cerebral ganglion. Hold the specimen (Fig. Explain its significance. This is a question and answer forum for students, teachers and general visitors for exchanging articles, answers and notes. A very complex study of this phenomenon evidenced the participation of high-voltage activated N-type calcium channels and the activation of noncapacitative calcium entry (NCCE). Many thanks also to all collaborators, especially to Dr. Dalia Gordon and Professor Michael Gurevitz from Tel Aviv University, Israel. It helped to discriminate between excitatory and depressant groups of toxins. In the presence of the toxin (10−8 M), unclustered, brief single channel openings in control (at −50 mV) were transformed into large, multistep amplitude bursting activity, separated by periods of silence. Such effect was limited after pretreatment with atropine. We will be providing unlimited waivers of publication charges for accepted research articles as well as case reports and case series related to COVID-19. In this article we will discuss about the dissection of cockroach. Oesophageal or pharyngeal or occipi­tal ganglion or corpora cardiacum: Situated on both sides of oesophagus slightly above the hypo-cerebral ganglion. TOS4. After one connective is desheathed, the giant axons become accessible for microelectrodes. In voltage-clamp experiments, short sodium current is prolonged during depolarizing voltage pulse (Figure 4(b)); potassium current is not modified. I. Huber, E. P. Masler, and B. R. Rao, Eds., Y. Pichon and J. Boistel, “Current-voltage relations in the isolated giant axon of the cockroach under voltage-clamp conditions,”, M. Pelhate and D. B. Sattelle, “Pharmacological properties of insect axons: a review,”, M. Pelhate, Y. Pichon, and D. J. Beadle, “Cockroach axons and cell bodies: electrical activity,” in, M. Pelhate and Y. Pichon, “Selective inhibition of potassium current in the giant axon of the cockroach,”, Y. Pichon and J. Boistel, “Microelectrode study of the resting and action potentials of the cockroach giant axon with special reference to the role played by the nerve sheath,”, E. Zlotkin, H. Rochat, Kopeyan, F. Miranda, and S. Lissitzky, “Purification and properties of the insect toxin from the venom of the scorpion, M. Pelhate and E. Zlotkin, “Actions of insect toxin and other toxins derived from the venom of the scorpion, D. Lester, P. Lazarovici, M. Pelhate, and E. Zlotkin, “Purification, characterization and action of two insect toxins from the venom of the scorpion, I. Karbat, F. Frolow, O. Froy et al., “Molecular basis of the high insecticidal potency of scorpion, D. Gordon, I. Karbat, N. Ilan et al., “The differential preference of scorpion, M. Gurevitz, I. Karbat, L. Cohen et al., “The insecticidal potential of scorpion, D. Gordon, “Sodium channels as targets for neurotoxins. As illustration, binding assays of many radiolabeled toxins have been successfully characterized with synaptosomes [24, 42, 43, 68]. Such axonal preparation exhibits simple bioelectrical properties; one type of voltage-dependent sodium and potassium (Kdr) channels can be found; they are responsible for generating short (0.5 ms) action potentials. Cockroach dissection - Nervous system - YouTube Cockroach Dissection Scientific Background One way to study an organism is to conduct a dissection. The synaptosomal preparation is easy to obtain and has been applied in several pharmacological tests. Cercal nerves X and XI are connected with giant interneurone dendritic tree by inhibitory and excitatory synapses, respectively, located in the last abdominal ganglion. There have also been cases of finding toxins of which the effects on axonal electrophysiological activity appear intermediate between those of excitatory and depressant groups, for instance: BcTx1 from East African scorpion Babycurus centrurimorphus [26] or BoT IT2, which, moreover, induced a new, unusual current with slow activation/deactivation kinetics [23, 24, 27]. Various types of receptors provide a very precise regulation of the neurons’ spontaneous activity and neurosecretory function. There were several experiments on DUM cells performed simultaneously with observations on an isolated axon. They appeared to be a new target for LqhαIT toxin, even more sensitive than the classical voltage-dependent Na channels in this preparation. Higher presynaptic activity results in the increased releasing of acetylcholine, however, it activates the negative feedback via presynaptic muscarinic receptors in turn and after several minutes of the toxin’s action, a decrease in postsynaptic events can be observed (Stankiewicz, personal observations). Unitary excitatory postsynaptic potentials (uEPSP) result from the stimulation of single hair mechanoreceptors covering the cercus; composed postsynaptic potentials (cEPSP) are the effect of cercal nerve XI electrical stimulation. Detach the glands and receptacles from the crop, separate the hypo-­pharynx from the specimen by cutting it at the base and the salivary apparatus is free (Fig. Stimulation of nerve X evokes inhibitory postsynaptic potentials (IPSP) via activation of GABA receptors [61]. The foregut extends in the form of the esophagus that has a thin wall(narrow) structure. Evoked responses are generated by electrical stimulation of the cercal nerve or direct mechanical stimulation of the cerci. Alpha-bungarotoxins and k-bungarotoxins completely blocked uEPSP and cEPSP at concentration of 10−7 M. All performed experimental protocols indicated that the toxins block neuronal postsynaptic nicotinic receptors and α-bungarotoxin was more effective [64, 65]. Two, short, wide tubes. Share Your PPT File. These are two solid nerves and run along the mid-ventral line of the thorax and abdomen. A round, thick walled muscular structure, posterior to the crop. Synaptic preparation from the cockroach represents a remarkably useful model for pharmacological studies, however, obtaining a high-quality stable preparation requires much experience. In the following paragraphs, various electrophysiological methods using cockroach preparations will be presented, along with their contribution to the development of toxinology. The field of toxinology owes much to the use of various nervous preparations obtained from this insect. Their applicability as bioinsecticides is now under consideration [12, 14, 15]. On the other hand, some arthropod neurotoxins show high selectivity to insect nervous system and they are considered as potent bioinsecticides. In the subsequent years, the story developed with the application of more and more modern methods and presently many details of arthropod toxins’ selectivity to insects are known [11–13]. The supra-oesophageal ganglion supplies the nerves to antennae and compound eyes. Pin down the head of the cockroach with ventral surface upward. The three thoracic ganglia and the last abdominal ganglion are large in size. The experiments performed with calcium imaging demonstrated the rise in the intracellular calcium concentration in the presence of toxin VII. For first year, we had dissection of cockroach and frog, which requires dissection box to carry out the operation.dissection box to carry out the operation. Although DUM neurons represent a much more complex model of bioelectrical activity than the axon, the observations were similar and the conclusions-compatible [25, 52]. 6.4). Paired, small in size, made up of a number of vesicles. Those from the abdominal ganglia send nerves to the structures in the abdominal region. In current-clamp, short action potentials are extended and transformed into plateau action potentials (Figure 4(a)). In toxicity tests, these toxins caused an immediate contraction paralysis of fly larvae and a quick excitatory “knock-down” effect on locusts [8, 10], which remains consistent with the observations done in electrophysiological experiments. An ill-defined chamber, bounded anteriorly by epipharynx and labrum; posteriorly by hypo-pharynx and labium; later­ally by two mandibles (Fig. 2018 Mar;11(2):185-195. doi: 10.1002/ase.1721. In such case, axon is left on its place in nervous chain, stays in contact with others axons and its microenvironment is much less changed compared to isolated axon. Give a transverse incision along the anterior border of the first thoracic segment and carefully remove the terga. Similar modification of axonal bioelectrical activity by Bot IT2 has been evidenced using a toxin from the venom of ant, Paraponera clavata, also tested on the cockroach isolated axon [28]. They and their collaborators isolated the genetic material responsible for the synthesis of toxins from the scorpion venom, they defined the cDNA sequence and developed artificial expression systems [33–36]. Cockroach Dissection Scientific Background One way to study an organism is to conduct a dissection. A dissection is a detailed investigation of that organism\u2019s organs and tissues. It has a long and coiled alimentary canaldivided into- foregut, midgut, and hindgut. This paper presents its role as a preparation in the development of toxinological studies in the following electrophysiological methods: double-oil-gap technique on isolated giant axon, patch-clamp on DUM (dorsal unpaired median) neurons, microelectrode technique in situ conditions on axon in connective and DUM neurons in ganglion, and single-fiber oil-gap technique on last abdominal ganglion synapse. The alimentary canal starts with the foregut which comprises the mouth and surrounded parts of the mouth. Foregut- It is also known as stomadaeum and is divided into buccal chamber, pharynx, oesophagus, crop, and gizzard. Review articles are excluded from this waiver policy. Scorpion alpha toxins bind to the receptor site 3 on sodium channel and inhibit the channel’s inactivation [12]. Cut the lateral membrane (pleura) between the terga and sterna of the thorax and abdomen with a pair of fine scissors. The long history of anti-insect scorpion toxins started with this technique, when in the 1970–80s the toxins purified from North African scorpions (Androctonus australis Hector—AaH IT and Buthus judaicus—Bj IT) by Professor Eliahu Zlotkin and collaborators from the Hebrew University of Jerusalem, Israel, and Faculté de Médecine, Marseille, France [8], were tested for the first time on cockroach isolated giant axon by electrophysiologist Professor Marcel Pelhate, at Angers University, France [9, 10]. The ovarioles unite to form the oviduct. Periplaneta americana represents an excellent model applied in different pharmacological methods, especially in electrophysiology, which plays a vital part in most of research activity in toxinology. Its effect on cockroach axon can also be described as intermediate between those of excitatory and depressant toxins. Pharmacological characteristics of alpha-like toxins are similar but not identical with those of alpha neurotoxins. It may be based on using natural and “artificial” preparations, as for example the transfected Xenopus oocytes. Fix the head of the speci­men by pinning through the mandibles. Arising from the brain two short and broad nerves run around the oesophagus to meet the sub-oeso­phageal ganglia (Fig. A narrow tube run­ning from the gizzard to the hind gut. The central nervous system of cockroach (Fig. Nervous system of Periplaneta americana cockroach is used in a wide range of pharmacological studies, including electrophysiological techniques. They have also been used in toxinological experiments. Zoology, Practicals, Dissection, Invertebrata, Nocturnal Insects, Cockroach. Lastly, the experiments performed on cockroach synaptic preparation contributed to studies which allowed to explain the mechanism of synergistic interaction between chemical neurotoxins: pyrethroids (permethrin) and carbamates (propoxur) [67]. Several toxins related to these groups were tested and their mode of action was determined using the single-fiber oil-gap method. Excretory System 10. Respiratory System 8. 6.5). Abstract The methods presented here demonstrate laboratory procedures for the dissection of four different regions of the central nervous system (CNS) from murine neonates for the isolation of glial subpopulations. Double-oil-gap technique permits, in current-clamp, to evoke large (up to 100 mV) axonal action potentials (Figure 2(b)(A)), to follow their evolution and to control the resting potential as well as the level of local response. Such effects were observed when the first anti-insect toxin LqhαIT from Leiurus quinquestriatus hebraeus scorpion venom was tested on cockroach isolated axon [32, 33]. After reading this essay you will learn about:- 1. What would be the consequences if there is no meiosis in organisms that reproduce sexually? A pair of connectives run backward from the ganglia and join the first thoracic ganglion, (prothoracic gang­lion). Passive characteristics of axonal membrane (resistance and capacitance) can be analyzed using longer (e.g., 5 ms) hyperpolarizing pulses (Figure 2(b)(B)); the application of long depolarizing pulses allows to observe delayed rectification (Figure 2(b)(B)). An important conclusion has been drawn from these studies that the reduction of the activity of NCCE may prove a useful strategy in the development of a drug for antienvenoming therapy [55]. Scorpion alpha toxin LqhαIT induced a substantial increase in the postsynaptic spontaneous activity, that is, in the frequency and the amplitude of uEPSP (Figures 6(b) and 6(c)), as well as in the amplitude and duration of cEPSP (Figure 6(d)). Biophysical principles of the nervous system function in insects are much the same as in mammals. In voltage-clamp configuration, depolarizing voltage pulses induce a short, inwardly directed, tetrodotoxin-sensitive sodium current and delayed, noninactivating outward potassium current (Figure 2(d)), most sensitive to 3,4 diaminopyridine [6]; hyperpolarizing pulses are used to record the leak current. Copyright © 2012 Maria Stankiewicz et al. They possess unmyelinated, very long (1.5–2 cm), large diameter (up to 50 μm) axons. It is formed by the fusion of two ganglia and located in the mid-ventral region of the head just ventral to the oesophagus. 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