Heinz Penzlin

Periviscerokinin (Pea-PVK): A novel myotropic neuropeptide from the perisympathetic organs of the American cockroach

A myotropic neuropeptide was isolated from extracts of 1000 abdominal perisympathetic organs of males of the cockroach, Periplaneta americana. This peptide, termed periviscerokinin, has excitatory actions on the hyperneural muscle of Periplaneta americana. After peptide sequence analysis and mass spectrometry, the structure of this peptide was confirmed by chemical synthesis and bioassay to be Gly-Ala-Ser-Gly-Leu-Ile-Pro-Val-Met-Arg-Asn-NH2. This sequence is different from the other known myotropic peptides in insects. The threshold concentration for stimulatory effects of the synthetic peptide on the isolated hyperneural muscle was about 10(-9) M, suggesting a physiological role as a neurohormone.

Isolation of Periviscerokinin-2 from the Abdominal Perisympathetic Organs of the American Cockroach, Periplaneta americana

Using the isolated hyperneural muscle as bioassay, a novel myotropin was isolated from the abdominal perisympathetic organs of Periplaneta americana. This is the second neuropeptide identified from insect perisympathetic organs. Peptide sequence analysis and mass spectrometry yielded the following structure: Gly-Ser-Ser-Ser-Gly-Leu-Ile-Ser-Met-Pro-Arg-Val-NH2. This peptide, named periviscerokinin-2, was confirmed to be amidated by chemical synthesis, bioassay, and comparison of retention times between native and synthetic peptides. A highly specific antiserum was used to determine sites of synthesis in the abdominal ganglia. Besides periviscerokinin-1, periviscerokinin-2 is the only putative myotropic neurohormone from the abdominal perisympathetic organs that is effective in the nanomolar range. This confirms the hypothesis that the neurohormonal system of the ventral nerve cord is remarkably different from that of the brain.

Isolation and structural elucidation of two pyrokinins from the retrocerebral complex of the American cockroach.

By monitoring the contractile activity of the hyperneural muscle of the American cockroach in vitro two peptides were isolated from the retrocerebral complex of the American cockroach. Three purification steps using reversed-phase high performance liquid chromatography on C-18 columns containing trifluoroacetic acid or heptafluorobutyric acid as organic modifiers were sufficient to achieve homogeneous peptide preparations. The structures of both peptides were elucidated by a combination of Edman degradation and mass spectrometry which yielded the following structures: His-Thr-Ala-Gly Phe-Ile-Pro-Arg-Leu-NH2 (Pea-PK-1) and Ser-Pro-Pro-Phe-Ala-Pro-Arg-Leu-NH2 (Pea-PK-2). The C-terminal sequence Phe-X-Pro-Arg-Leu-NH2 characterized the peptides as members of the insect pyrokinin family. The synthetic peptides were shown to have the same retention times as the natural peptides. The occurrence of both peptides in the retrocerebral complex suggests a physiological role as neurohormones. The effects of the synthetic pyrokinis were clearly distinguishable in their actions on the hyperneural muscle. Regarding the threshold concentrations, Pea-PK-2 was only 0.3% as active as Pea-PK-1.

Isolation and structural elucidation of eight kinins from the retrocerebral complex of the American cockroach, Periplaneta americana

By monitoring the contractile activity of the hindgut of the American cockroach in vitro eight myotropic neuropeptides were isolated from the retrocerebral complex of the American cockroach. Peptide sequence analysis and mass spectrometry yielded the following structures: Arg- Pro-Ser-Phe-Asn-Ser-Trp-Gly-NH2 (Pea-K-1), Asp-Ala-Ser-Phe-Ser-Ser-Trp-Gly-NH2 (Pea-K-2), Asp-Pro-Ser-Phe-Asn-Ser-Trp-Gly-NH2 (Pea-K-3), Gly-Ala-Gln-Phe-Ser-Ser-Trp-Gly-NH2 (Pea-K-4), Ser-Pro-Ala-Phe-Asn-Ser-Trp-Gly-NH2 (Pea-K-5), Asp-Pro-Ala-Phe-Ser-Ser-Trp-Gly-NH2 (Lem-K-7), Gly-Ala-Asp-Phe-Tyr-Ser-Trp-Gly-NH2 (Lem-K-8) and Ala-Phe-Ser-Ser-Trp-Gly-NH2 (Lom-K). The C-terminal sequence Phe-X-Ser-Trp-Gly-NH2 characterized the peptides as members of the insect kinin family. All structures were confirmed by comparison of retention times between synthetic and natural peptides. The threshold concentration for stimulatory effects of the synthetic peptides on the isolated hindgut was about 10(-9) M and there was no significant difference measured between the different kinin forms. These neuropeptides are the first members of the insect kinin-family isolated from the American cockroach. Their occurrence in the retrocerebral complex suggests a physiological role as neurohormone.

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.

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 Ringers solution has no significant effect on the learning process in the lifting task. In the lowering task, however, Ringers solution impairs the learning ability considerably. Crude extract of corpora cardiaca strongly inhibits the learning of both tasks.

The effect of diflubenzuron (Dimilin) on the ecdysteroid titer and neuronal activity of Periplaneta americana (L.)

Abstract The insecticide diflubenzuron (DFB) affected the larval-adult development of Periplaneta americana when administered by feeding or injection. Usually, the larvae died during adult ecdysis due to the inability to split their larval cuticle. In other cases, they did ecdyse partially or completely, but died during the first day after emergence. The duration of the larval instar after DFB treatment was prolonged. By radioimmunoassay the in vitro ecdysteroid production of the prothoracic gland as well as the ecdysteroid titer of the hemolymph in different periods of the last larval instar of P. americana was determined. It was shown that the first increase of the ecdysteroid production of the molting gland during the instar was unaffected by DFB in contrast to the second peak in the ecdysteroid production, which was missing. The dose-response relationship of the effect of DFB and dimethylsulfoxide (DMSO) as the solvent of DFB on the spontaneous neuronal activity of the frontal ganglion was studied in vitro. Administration of these substances resulted in a high stimulation of the spontaneous burst activity coming from the frontal ganglion and going via nervus connectivus to the somata in the protocerebrum. The action of DFB and DMSO seems to be a direct effect on the cholin-sensitive sites at synaptic regions in addition to the inhibition of the acetylcholinesterase (EC 3.1.1.7).