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PubMed1999

The possibility of testing antiparasitical substances using the cockroach as a model.

Products used against parasitic arthropods must be tested for their efficacy. Because the supply of parasitic organisms for such tests may be a limiting factor, it is best to conduct these tests with a model organism. We show that the cockroach is a suitable model organism because it is easy to keep and to breed. Moreover, all stages of these animals are more resistant than, or at least as resistant as, the corresponding parasites, so the results with cockroaches can also be taken as valid for ectoparasites.

Animals↗
PubMed2002

Neuronal co-localization of different isoforms of tachykinin-related peptides (LemTRPs) in the cockroach brain.

Seven isoforms of tachykinin-related peptides (TRPs) have been isolated from the brain of the cockroach Leucophaea maderae. These peptides (LemTRP-1, 2, and 5-9) share the C-terminal sequence GFX(1)GX(2)Ramide (where X(1) and X(2) are variable residues). In order to determine the neuronal distribution of several of these LemTRP isoforms, we raised antisera to their variable N-termini. Antisera to LemTRP-1, 2, 3, 7, and 8 were utilized for immunocytochemistry on cryostat sections of the L. maderae brain. As expected, the gut peptide LemTRP-3 was not detected in the brain, and the antisera to LemTRP-1, 2, and 7 labeled the same sets of neurons in different regions of the brain. These neurons could also be labeled with antisera raised to the more conserved C-termini of LemTRP-1 and the locust TRP LomTK-I. The antiserum to LemTRP-8 predominantly labeled a set of neurons distinct from that seen with any other N- or C-terminus-directed antisera, suggesting that it recognizes epitope(s) other than known insect TRPs. Our findings indicate that at least three of the LemTRPs are always co-localized in neurons of the L. maderae brain. We have also been able to show that LemTRP-2, which is an N-terminally extended form (17-mere) of LemTRP-1 with a dibasic putative cleavage site, is transported throughout the processes of the neurons in the same manner as LemTRP-1 and 7. Thus, LemTRP-2 may be released with the other shorter LemTRPs. This is the first investigation of LemTRP distribution in the cockroach central nervous system utilizing antisera to native peptides.

Amino Acid Sequence↗
PubMed2003

Ultrastructure of pigment-dispersing hormone-immunoreactive neurons in a three-dimensional model of the accessory medulla of the cockroach Leucophaea maderae.

Locomotor activity rhythms of the cockroach Leucophaea maderae are orchestrated by two bilaterally symmetric, mutually coupled, circadian pacemakers. They lie in the optic lobes of the brain and are confined to the accessory medulla (AMe), ventro-medially to the medulla. The AMe is innervated by approximately 12 pigment-dispersing hormone (PDH)-immunoreactive anterior medulla neurons (PDHMe), which are circadian pacemaker candidates in the fruitfly and the cockroach. We have developed a three-dimensional computer model of the AMe and associated structures as a framework for neuroanatomical studies. Our greatly improved understanding of this structure in space has allowed us further to subdivide the anterior PDHMe into three subgroups, i.e., large, medium-sized, and small anterior PDHMe. The synaptic connections of two of these subgroups have been examined within subcompartments of the AMe by light and electron microscopy. The large, intensely staining, anterior PDHMe contain medium-sized dense-core vesicles and form input and output synapses with profiles densely filled with clear vesicles primarily in the anterior and shell neuropil of the AMe. The medium-sized anterior PDHMe contain large dense-core vesicles and constitute input and output synapses either with profiles being densely filled with clear vesicles, or with profiles containing granular dense-core vesicles. The small, weakly staining anterior PDHMe belong to a morphological group different from the large and medium-sized PDHMe and cannot be further identified at the electron-microscopic level because of their weak PDH immunoreactivity.

Animals↗
PubMed2004

Pigment-dispersing hormone (PDH)-immunoreactive neurons form a direct coupling pathway between the bilaterally symmetric circadian pacemakers of the cockroach Leucophaea maderae.

Circadian locomotor activity rhythms of the cockroach Leucophaea maderae are driven by two bilaterally paired and mutually coupled pacemakers that reside in the optic lobes of the brain. Transplantation studies have shown that this circadian pacemaker is located in the accessory medulla (AMe), a small neuropil of the medulla of the optic lobe. The AMe is densely innervated by about 12 anterior pigment-dispersing-hormone-immunoreactive (PDH-ir) medulla (PDHMe) neurons. PDH-ir neurons are circadian pacemaker candidates in the fruitfly and cockroach. A subpopulation of these neurons also appears to connect both optic lobes and may constitute at least one of the circadian coupling pathways. To determine whether PDHMe neurons directly connect both accessory medullae, we injected rhodamine-labeled dextran as neuronal tracer into one AMe and performed PDH immunocytochemistry. Double-labeled fibers in the anterior, shell, and internodular neuropil of the AMe contralaterally to the injection site showed that PDH-ir fibers directly connect both accessory medullae. This connection is formed by three anterior PDHMe neurons of each optic lobe, which, thus, fulfill morphological criteria for a direct circadian coupling pathway. Our double-label studies also showed that all except one of the midbrain projection areas of anterior PDHMe neurons were innervated ipsilaterally and contralaterally. Thus, anterior PDHMe neurons seem to play multiple roles in generating circadian rhythms. They also deliver timing information output and perform mutual pacemaker coupling in L. maderae.

Animals↗
PubMed2006

Orcokinin immunoreactivity in the accessory medulla of the cockroach Leucophaea maderae.

The accessory medulla is the master circadian clock in the brain of the cockroach Leucophaea maderae and controls circadian locomotor activity. Previous studies have demonstrated that a variety of neuropeptides are prominent neuromediators in this brain area. Recently, members of the orcokinin family of crustacean neuropeptides have been identified in several insect species and shown to be widely distributed in the brain, including the accessory medulla. To investigate the possible involvement of orcokinins in circadian clock function, we have analyzed the distribution of orcokinin immunostaining in the accessory medulla of L. maderae in detail. The accessory medulla is densely innervated by approximately 30 orcokinin-immunoreactive neurons with cell bodies distributed in five of six established cell groups in the accessory medulla. Immunostaining is particularly prominent in three ventromedian neurons. These neurons have processes in a median layer of the medulla and in the internodular neuropil of the accessory medulla and send axonal fibers via the posterior optic commissure to their contralateral counterparts. Double-labeling experiments have revealed the colocalization of orcokinin immunostaining with immunoreactivity for pigment-dispersing hormone, FMRFamide, Mas-allatotropin, and gamma-aminobutyric acid in two cell groups of the accessory medulla, but not in the ventromedian neurons or in the anterior and posterior optic commissure. Immunostaining in the ventromedian neurons suggests that orcokinin-related peptides play a role in the heterolateral transmission of photic input to the pacemaker and/or in the coupling of the bilateral pacemakers of the cockroach.

Animals↗
PubMed1997

Improved conditions for culture of biosynthetically active cockroach corpora allata.

Currently, short-term culture of insect corpora allata is most often performed in TC199. We now show that L-15B, a medium widely used in arthropod tissue culture, is superior to TC199 for both short- and long-term culture of cockroach corpora allata. In 3-h and 48-h incubations, juvenile hormone biosynthesis by corpora allata from Diploptera punctata was significantly higher in L-15B than in TC199. In addition, in both media, corpora allata activity was significantly improved by flotation of glands at the medium surface. Characteristics of L-15B responsible for its superiority were examined by comparison of gland activities in several TC199 formulations that had been modified in different ways to be more similar to L-15B. Adjusting the osmotic pressure of TC199 (288 mOsm/l) to near that of L-15B (362 mOsm/l) and D. punctata hemolymph (360 mOsm/l) significantly improved gland activity during the second 12 h of a 36-h incubation. Increasing the concentrations of amino acids, sugars, and organic acids in TC199 to the same levels as in L-15B significantly improved gland activity during both the second and third 12-h intervals of a 36-h incubation. These results suggest that L-15B is superior to TC199 because L-15B is isoosmotic with D. punctata hemolymph and because L-15B, like cockroach hemolymph, contains a high level of organic constituents. It is therefore more appropriate to use L-15B than TC199 for short-term in vitro assays of juvenile hormone biosynthesis and for extended corpora allata culture.

Animals↗
PubMed1987

De novo biosynthesis of polyunsaturated fatty acids in the cockroach Periplaneta americana.

The de novo biosynthesis of 6,9,12-linolenic acid, 11,14-eicosadienoic acid, 5,11,14-eicosatrienoic acid, and arachidonic acid was demonstrated in adult female cockroaches, Periplaneta americana. These four polyunsaturated fatty acids (PUFA) were present primarily in the phospholipid (PL) fraction of both males and females. They were purified by AgNO3 thin-layer chromatography and high pressure liquid chromatography. The double bond positions of the major isomer of eicosatrienoic acid were shown to be at the delta 5,11,14 positions by gas chromatography-mass spectrometry (GC-MS) of both methoxy and epoxide derivatives and gas-liquid chromatography (GLC) and GC-MS of ozonolysis products. The other PUFAs cochromatographed with standards on both packed and capillary GLC columns. The in vivo incorporation of [1-14C]acetate into 5,11,14-eicosatrienoic acid, 11,14-eicosadienoic acid, 6,9,12-linolenic acid, and arachidonic acid was demonstrated by radio-GLC and radio-HPLC and for 5,11,14-eicosatrienoic acid by radio-GLC of ozonolysis products. The latter technique clearly demonstrated that the entire eicosatrienoic acid molecule was labeled. Thoracic tissue contained the highest amount of radiolabeled 5,11,14-eicosatrienoic acid (1.6% of total radioactivity incorporated into PL) while radiolabeled 11,14-eicosadienoic acid was found primarily in abdominal epidermal tissue (2% of total radioactivity incorporated into PL). Radiolabeled arachidonic and 6,9,12-linolenic acids comprised 0.1 and 0.02%, respectively, of the total radioactivity in the PL fraction. These data document the de novo biosynthesis of di-, tri-, and tetraunsaturated fatty acids in the American cockroach, and indicate that this animal can desaturate on both sides of the delta 9 double bond of oleic acid.

Abdomen↗
PubMed1975

Localization of the cockroach optic lobe circadian pacemaker with microlesions.

Electrolytic microlesions were made at various sites in optic lobes of cockroaches Leucophaea maderae, and animals were assayed for circadian locomotor rhythms. Lesions placed in the lamina or medulla disrupted rhythmicity in only a few animals (3 out of 55). Lesions placed in or near the lobula produced a greater fraction of arrhythmic roaches (18 out of 45). Over half of these lesions either included or were centered in the cell body regions bordering the second optic chiasm and the lobula. The results suggest that the cell bodies and not the neuropile areas of the lobe are the crucial elements of the clock driving the cockroach's circadian activity rhythm.

Animals↗
PubMed1977

The path and rate of growth of regenerating motor neurons in the cockroach.

Nerve root 5 that supplies the coxal depressor muscles from the metathoracic ganglion in the cockroach was crushed. Regeneration of the motor neurons was studied by cutting the nerve at several distances from the crush point and introducing cobalt chloride into the cut end. The operation was followed by a lag period of about 13 days after which the axons regenerated at a rate of 0.9 mm per day. After regeneration had been completed a pattern of axon distribution was established among the nerve branches that was very similar to that found in intact, normal cockroaches. This pattern was established through an apparent directed growth of certain axons from identified cells into branch 5rl, their normal pathway. However, at the same time, radom or increased branching of other unidentified motor neurons produced some errors in the distribution of axons among the nerve branches. Eventually these errors were corrected by the degeneration of neuronal processes that did not synapse with correct target muscles. These results demonstrate the requirement for a highly specific intercellular recognition process between individual, identified motor neurons and the appropriate muscles they innervate in order to reform the original innervation pattern during regneration.

Animals↗
PubMed1985

Analyzing cockroach escape behavior with lesions of individual giant interneurons.

Individual giant interneurons (GIs) in the ventral nerve cord of the cockroach, Periplaneta americana, were lesioned by intracellular injection of proteolytic enzymes (pronase). This was accomplished with minimal dissection, so that the wind-evoked escape behavior of the animals could be studied following the lesion. Unilateral lesions of GI-2 had no obvious effect on escape behavior, but unilateral removal of GI-1, as well as combined unilateral lesions of GIs 1 and 2, influenced the direction of an animal's initial turning movement in response to a wind puff. These results support the hypothesis that GIs play a role in initiating and guiding the directional, wind-evoked escape response of the cockroach.

Animals↗
PubMed1988

Escape responses following elimination of the giant interneuron pathway in the cockroach, Periplaneta americana.

The entire set of giant interneurons (GIs) in the nerve cord of the cockroach, Periplaneta americana, was ablated using either electrolytic or surgical techniques. All animals with these lesions were capable of turning and running away from standard wind puffs. However, all animals responded much less frequently to standard wind stimuli following lesion, and the latency of their responses was significantly increased. These results are discussed in terms of a GI role in extremely short latency escape responses, and the idea that non-GI pathways, perhaps associated with head sensory structures, need to be considered in the normal control of escape in the cockroach.

Animals↗
PubMed1985

Plasticity in the cockroach neuromuscular system.

The retrograde transport of wheat germ agglutinin-conjugated horseradish peroxidase extracellularly injected into a leg muscle was used to identify the regenerating cockroach motor neurons that have grown an axonal branch into that muscle. At least 66% of the animals with crushed nerve roots eventually reform the original innervation pattern of this muscle with no mistakes. In spite of this apparent specificity the cockroach neuromuscular system can express plasticity as evidenced by the correction of mistakes made at early stages of regeneration. These mistakes are corrected through elimination during the time interval between 40 and 60 days after nerve crush. In addition, when the distal segments of the leg are removed, thus depriving some motor neurons of their normal target muscles, many of them form stable inappropriate axonal branches in denervated as well as fully innervated muscles. These observations are discussed in terms of possible mechanisms responsible for the specificity of the cellular interactions and in terms of their relevance to understanding the development of vertebrate neuromuscular systems.

Axons↗
PubMed1984

Trehalases from the male accessory glands of the American Cockroach: developmental changes and the hormonal regulation of the enzymes.

Two types of trehalases, designated CM-I and CM-II, were detected in the male accessory gland of the American cockroach and they could be separated by CM-cellulose chromatography (S. Y. Takahashi, S. Higashi, S. Minoshima, M. Ogiso, and K. Hanaoka, 1980, Int. J. Invert. Reprod. 2, 373-381). Trehalase activity in the gland showed a rapid increase after adult emergence. The relative activities of the two enzymes were followed separately during adult development. The appearance of CM-II preceded that of CM-I during adult development. Allatectomy and decapitation of newly molted adults resulted in inhibition of the increase of enzyme activity, and in the allatectomized cockroach, CM-II, which is the major enzyme activity in the gland, was missing. Implantation of the corpora allata as well as application of JH-III and the JH analogs isopropyl-11-methoxy-3,7,11-trimethyl-2,4,-dodecadienoate (ZR-515) and 6,7-epoxy-1-(p-ethylphenoxy)-3,7-dimethyl-2-octene (R-20458) into the decapitated animals restored the enzyme activities. The data suggested that trehalase in the male accessory gland was under the control of the corpora allata. The regulation of trehalase activity in the male accessory gland was discussed with respect to its function.

Animals↗
PubMed1989

The hypertrehalosaemic peptides of cockroaches: a phylogenetic study.

Hypertrehalosaemic peptides from the corpora cardiaca of 14 different species were compared with respect to phylogenetic relationships within the insect suborder Blattaria (cockroaches). Gland extracts from members of the family Blattidae (Periplaneta americana, P. brunnea, P. australasiae, P. fuliginosa, and Blatta orientalis) contain two hypertrehalosaemic octapeptides with identical properties to the recently sequenced peptides M I and M II from the American cockroach, whereas corpora cardiaca from members of the families Blaberidae and Blattellidae (Nauphoeta cinerea, Leucophaea maderae, Blaberus discoidalis, B. trapezoideus, Diploptera punctata, and Gromphadorhina portentosa) possess one hypertrehalosaemic decapeptide with identical properties as the peptide recently sequenced from B. discoidalis and N. cinerea. A member of the family of Polyphagidae (Polyphaga aegyptiaca), placed at the origin of the phyletic tree of Blattaria, has two hypertrehalosaemic factors in its corpus cardiacum which are each different from M I, M II, and HTH.

Amino Acid Sequence↗
PubMed1992

Primary structures of the hypertrehalosemic peptides from corpora cardiaca of the primitive cockroach Polyphaga aegyptiaca.

Two hypertrehalosemic neuropeptides from the corpus cardiacum of the cockroach Polyphaga aegyptiaca were isolated by reversed-phase high-performance liquid chromatography, and their primary structures were determined by pulsed-liquid phase sequencing employing Edman chemistry, after enzymically deblocking the N-terminal pyroglutamate residue. As neither peptide was cleaved by carboxypeptidase, the C-terminus of each peptide was also blocked. Both peptides were found to be uncharged octapeptides with the sequences: Peptide 1: pGlu-Leu-Asn-Phe-Ser-Pro-Asn-Trp-NH2; and peptide 2: pGlu-Ile-Thr-Phe-Thr-Pro-Asn-Trp-NH2. Both peptides are clearly defined as members of the adipokinetic hormone/red pigment-concentrating hormone family of peptides. Whereas peptide 1 is identical in structure to the previously sequenced hypertrehalosemic neuropeptide from tenebrionid beetles (and is therefore designated the acronym Tem-HrTH), peptide 2 is a novel peptide and is designated the acronym Poa-HrTH. Both synthetic peptides caused an increase in the hemolymph carbohydrate concentration in P. aegyptiaca, specifically changing the trehalose concentration. The novel peptide Poa-HrTH was not very potent in elevating blood carbohydrates in the American cockroach.

Amino Acid Sequence↗
PubMed1987

Pharmacological characterisation of the dopamine-sensitive adenylate cyclase in cockroach brain: evidence for a distinct dopamine receptor.

Dopamine increases cyclic AMP production in crude membrane preparations of cockroach brain with plateaus in cyclic AMP production occurring between 1-10 microM and at 10 mM. Maximal production of cyclic AMP is 2.25 fold greater than that of control values. Octopamine also increases cyclic AMP production with a Ka of 1.4 microM and maximal production 3.5 fold greater than that of control. 5-Hydroxytryptamine does not increase cyclic AMP production. The effects of octopamine and dopamine are fully additive. The vertebrate dopamine agonists ADTN and epinine stimulate the dopamine-sensitive adenylate cyclase (AC) with Ka values of 4.5 and 0.6 microM respectively and with maximal effectiveness 1.7 fold greater than that of control. The selective D2-dopamine agonist LY-171555 stimulates cyclic AMP production to a similar extent with a Ka of 50 microM. Other dopamine agonists (apomorphine, SKF-82526, SKF-38393) have no stimulatory effects. The octopamine-sensitive AC is inhibited by a variety of antagonists known to affect octopamine and dopamine receptors, with the following order of potency: mianserin greater than phentolamine greater than cyproheptadine greater than piflutixol greater than cis-flupentixol greater than SCH-23390 greater than (+)-butaclamol greater than SKF-83566 greater than SCH-23388 greater than sulpiride greater than spiperone greater than haloperidol. The dopamine-sensitive AC is inhibited by the same compounds with the following order of potency: piflutixol greater than cis-flupentixol greater than (+)-butaclamol greater than spiperone greater than or equal to SCH-23390 greater than cyproheptadine greater than SKF-83566 greater than SCH 23388 greater than mianserin greater than phentolamine greater than sulpiride greater than haloperidol. With the exception of mianserin, 3H-piflutixol is displaced from brain membranes by dopamine antagonists with an order of potency similar to that observed for the inhibition of dopamine-sensitive AC. The results indicate that the octopamine- and dopamine-sensitive AC in cockroach brain can be distinguished pharmacologically and the dopamine receptors coupled to AC have pharmacological characteristics distinct from vertebrate D1- and D2-dopamine receptors.

Adenylyl Cyclases↗
PubMed1985

A comparison of shock avoidance learning in headless cockroaches in leg-lifting and lowering task.

Headless cockroaches were trained either to lift or to lower the prothoracic leg to avoid electric shocks. The learning process in both cases was very distinct. However, the learning performance in the lifting was better than in the lowering task. The reason for this poorer learning in the lowering task is due to the unconditioned leg raising movement of the animals to electric shocks. This unconditioned response affects the headless cockroaches avoid shocks in the lifting task by escape learning, whereas they avoid shocks in the lowering task by true avoidance learning. Injection of 30 microliter of Ringer's solution has no significant effect on the learning process in the lifting task. In the lowering task, however, Ringer's solution impairs the learning ability considerably. Crude extract of corpora cardiaca strongly inhibits the learning of both tasks.

Animals↗
PubMed1987

Effect of morphine and naloxone on shock avoidance learning in headless cockroaches (Periplaneta americana L.).

Systemic administration of high doses of morphine (56 micrograms morphine/g body weight) or of naloxone (54 micrograms naloxone/g body weight) results in a significant improvement of shock avoidance behavior in headless cockroaches. In both cases the learning parameter stimulation time (time during which an animal receives shocks) is significantly decreased. The behavioral parameters, stimulation rate (activity) and mean stimulation duration (shock responsiveness), as the two factors of the stimulation time considered individually, do not significantly change with morphine. Only the responses of both parameters together give the significant decrease of stimulation time (improvement of shock avoidance behavior) as mentioned above. However, the administration of naloxone causes a significant shortening of the mean stimulation duration indicating that these animals avoid the shocks by learning to lift their legs more quickly out of electrified saline in order to terminate the shocks (escape learning). Combined administration of both drugs together also causes a significant improvement of the leg-lifting response even at lower doses (morphine: 0.56 microgram plus naloxone: 0.54 microgram/g; morphine: 0.0056 microgram/g plus naloxone: 0.0054 microgram/g). Although the various effective doses of these drugs administered in combination have similar effects on the stimulation time, they affect the behavioral parameters in different ways. These different actions on activity and shock responsiveness as well as the efficacy of doses smaller than those known from vertebrates are discussed as behavioral evidence of opiate receptors in the cockroach.

Animals↗