Molluscan Cardioexcitatory Neuropeptide Research Articles

Characteristics of an Aminopeptidase from Japanese Cedar (Cryptomeria japonica) Pollen

A peptidase from Japanese cedar pollen, Jc-peptidase, was clarified to preferentially hydrolyze an MCA substrate of Phe-MCA (L-phenylalanyl-4-methylcoumaryl-7-amide). This study examined substrate specificities of Jc-peptidase using oligopeptides. Jc-peptidase hydrolyzed Phe-Phe and Tyr-Phe effectively and hydrolyzed Leu-Phe, Met-Phe, and Arg-Phe moderately. Other substrates such as Ala-Phe, Asp-Phe, and Pro-Phe were not hydrolyzed with the peptidase. Results obtained with a series of aminoacyl-Phe peptides were compatible with the facts obtained for MCA substrates except for Arg-MCA. Effects of amino acid residues in the P1' position were also examined using Phe-amino acids. An N-terminal phenylalanine residue was actually released from bioactive peptides such as molluscan cardioexcitatory neuropeptide (FMRF-NH(2)). Because the activity was inhibited with Zn(2+) and EDTA, Jc-peptidase was inferred to belong to the metalloproteases. The N-terminal amino acid sequence was determined to be APIGVQLEIEENYVHMYNGF and an internal sequence to be EIFAATFNVDEETEA, but no homology with other proteins was found.

Supraspinal administration of [D-Met 2]FMRFamide produces a naloxone-sensitive increase in heart rate in unrestrained spontaneously hypertensive rats

Intracerebroventricular (i.c.v.) administration of either Phe-Met-Arg-Phe-NH 2 (FMRFamide; molluscan cardioexcitatory neuropeptide; 3–30 μg) or the FMRFamide analog Phe-D-Met-Arg-Phe-NH 2 ([D-Met 2]FMRFamide; 15 μg) to conscious unrestrained spontaneously hypertensive rats (SHR) produced a relatively long lasting (> 1 h) increase in heart rate. The increase in heart rate produced by [D-Met 2]FMRFamide was attenuated by i.c.v. injection of the opiate antagonist naloxone (2 μg). These results extend to a second endpoint an apparent opioid agonist-like (naloxone-reversible) action of [D-Met 2]FMRFamide.

Two subpopulations of ganglion cells in hydra are distinguished by an immunogold-double-labeling technique

In the primitive nervous system of Hydra sensory cells and ganglion cells are easily distinguished by their location and distribution, but distinguishing subpopulations within these two groups of neurons is difficult. Both sensory cells and ganglion cells in Hydra and other coelenterates can be immunocytochemically stained with antisera against the molluscan cardio-excitatory neuropeptide FMRFamide as well as other peptides with the general structure < Glu... Arg-X-NH2, where X is an aromatic amino acid. Arg-Phe-amide (RFamide)-like peptides have be e n localized in densecored vesicles of ganglion cells in the peduncle of Hydra. but to date there is no ultrastructural evidence to distinguish between subpopulations of ganglion cells. Therefore, we used an immunogold-double-labeling procedure for electron microscopy with antisera to FMRFamide and RGamide (Arg-Gly-amide) to determine if these two peptides characterized different types of ganglion cells in the peduncle and lower part of the hypostome. In addition, we tested for colocalization of RFamide- and Antho-RFamide-like material in the same neuronal dense-cored vesicles.

Action of serine carboxypeptidase from Aspergillus saitoi on carboxyterminal amidated peptides.

The specificity of the serine carboxypeptidase from Aspergillus saitoi (EC 3.4.16.1) was investigated on carboxyterminal amidated peptides such as benzyloxycarbonyl(Z)-Ala-Phe-NH2, gastrin-related peptide, molluscan cardioexcitatory neuropeptide, eledoisin-related peptide, and (D-Ala2, Met5)-enkephalinamide. The enzyme did not hydrolyze Z-Ala-Phe-NH2. It acted only as a carboxyamidase for the carboxyterminal peptide bond of gastrin-related peptide and as an amidase for enkephalinamide. The enzyme had both carboxyamidase activity and amidase activity for molluscan cardioexcitatory neuropeptide and eledoisin-related peptide. Whether the enzyme acts as an amidase or carboxyamidase, the hydrophobicity of P3 and P4 positions of the substrate may be important. After removal of the carboxyterminal amino acid amide and/or ammonia, the enzyme catalyzed the sequential liberation of the amino acid from the carboxyterminus of the substrate.

CCK 8 analgesia and hyperalgesia after intrathecal administration in the rat: Comparison with CCK-related peptides

The C-terminal octapeptide of cholecystokinin (CCK 8) was administered intrathecally to rats. Doses in the nanogram range produced weak but significant antinociception in the paw pressure test five minutes after injection whereas microgram doses of CCK 8 produced hyperalgesia. The CCK 8-induced analgesia or hyperalgesia was not seen in the tail flick test and was not associated with motor incapacitation or any other noticeable side effects. The C-terminal tetrapeptide of CCK (CCK 4) and pentagastrin were found to be ineffective in all tests but caerulein and molluscan cardioexcitatory neuropeptide (FMRF-amide), like CCK 8, produced antinociception in the paw pressure test.

Diversity in neurohaemal organs for homologous neurosecretory cells in different insect species as demonstrated by immunocytochemistry with an antiserum to molluscan cardioexcitatory peptide

Neurosecretory cells were histochemically identified in the thoracic ganglia of the Colorado potato beetle and Drosophila melanogaster with an antiserum to the molluscan cardioexcitatory neuropeptide FMRFamide (Phe-Met-Arg-Phe-NH 2). These cells and the FMRFamide-immunoreactive neurosecretory cells identified in a locust are most likely homologous. The neurosecretory axon terminals in the 3 species use different neurohaemal organs. Such a structural diversity in release sites of apparently homologous neurosecretory cells in different species may account for the large structural variability in neurohaemal organs in insects in general.

A comparison of the interaction of glucagon, human parathyroid hormone-(1-34)-peptide and calcitonin with dimyristoylphosphatidylglycerol and with dimyristoylphosphatidylcholine

The interaction of glucagon, human parathyroid hormone-(1–34)-peptide and salmon calcitonin with dimyristoylphosphatidylglycerol (DMPG) and with dimyristoylphosphatidylcholine (DMPC) was studied as a function of pH and temperature. The effect of lipid on the secondary structure of the peptide was assessed by circular dichroism and the effect of the peptide on the phase transition properties of the lipid was studied using differential scanning calorimetry. Some peptides interact more strongly with anionic than with zwitterionic phospholipids. This does not require an overall positive charge on the peptide. Increased thermal stability is observed in complexes formed between cationic peptides and anionic lipids. Particularly marked effects of glucagon and human parathyroid hormone-(1–34)-peptide on the phase transition properties of DMPG at pH 5 have been observed. The transition temperature is raised over 10°C at a lipid/peptide molar ratio of less than 30:1 and the transition enthalpy is increased over 2-fold. These effects do not occur with any basic peptide and were not observed with metorphinamide, molluscan cardioexcitatory neuropeptide or myelin basic protein. The results demonstrate that certain peptides can affect the phase transition properties of lipids in a manner similar to divalent cations. The overall hydrophobicities of these peptides can be evaluated by their partitioning between aqueous and organic solvents. None of the above three peptide hormones partition into the organic phase. However, a closely related peptide, human calcitonin, does exhibit substantial partitioning into the organic phase. Nevertheless, human calcitonin has a weaker interaction with both DMPC and DMPG than does salmon calcitonin. The effects of human calcitonin on the phase transition of DMPC are qualitatively different from those of salmon calcitonin in that the human form more readily eliminates the pretransition but causes less change in the main transition. Like overall charge, overall hydrophobicity is not an overwhelming factor in determining the ability of peptides to interact with phospholipids but rather more specific interactions are required for strong complexes to form.

Multiple actions of a molluscan cardioexcitatory neuropeptide and related peptides on identified Helix neurones.

The effects of the molluscan neuropeptide Phe-Met-Arg-Phe-NH2 (FMRF amide) and related peptides (Price & Greenberg, 1977) were tested on Helix aspersa neurones. Ionophoretic application of FMRFamide depolarized and excited some neurones, but hyperpolarized and inhibited others. In some neurones the sign of the response was dependent on the membrane potential. Two responses resulted from an increase in membrane conductance, a depolarizing response mediated mainly by an increase in Na+ ion permeability, and a hyperpolarizing response mediated by an increase in K+ ion permeability. In the C1 neurone a voltage-dependent response was observed, which only occurred when the neurone was depolarized from its resting level. This response was recorded as an inward current during voltage clamp and resulted from a decrease in K current(s), possibly Ca-activated K current. More than one response may occur in a single neurone. In the C1 neurone, the K-mediated hyperpolarization occurred as well as the voltage-dependent response, while the depolarization seen in the F2 neurone was a combination of an increase in Na conductance and an increase in K conductance.

Histochemical localization of FMRFamide-like immunoreactivity in the rat brain

Molluscan cardioexcitatory neuropeptide or FMRFamide is present in the invertebrate central nervous system (CNS) and FMRFamide like peptide has been demonstrated in the mammalian CNS. In this study, the distribution of FMRFamide immunoreactivity was studied in rat brain using the indirect immunofluorescent method. The highest number of FMRFamide staining cell bodies was found in the nucleus (n) arcuatus. N. paraventricularis, n. hypothalamus, n. ventromedialis, n. dorsomedialis and n. tractus solitarii also contained high numbers. FMRFamide positive nerve fibers and terminals were widely distributed. The septal complex contained high densities, especially in n. interstitialis striae terminalis. N. paraventricularis hypothalami, n. paraventricularis, n. hypothalamicus, n. ventromedialis and n. dorsomedialis showed a high to very high degree of immunoreactivity. In myelencephalon, n. tractus solitarii had the densest innervation. Spinal cord had a dense band of FMRFamide positive fibers in lamina I and II of the dorsal horn. The present findings support a neurotransmitter role for a FMRFamide like peptide in the mammalian brain, possibly related to endocrine and autonomic regulation as well as pain modulation.