A dense core vesicle protein is restricted to the cortex of granules in the exocrine atrial gland of Aplysia california.

We have generated a monoclonal antibody (mAb) 5E10 which recognizes an antigen localized to dense core vesicles (DCVs) in the atrial gland of Aplysia californica. mAb5E10 immunoprecipitates an abundant 57-kDa glycoprotein (atrial gland granule-specific antigen, AGSA) which is a soluble component of atrial gland DCVs. Electron microscopy reveals that AGSA immunoreactivity is restricted to the region between the dense core, which contains neuropeptide immunoreactivity, and the membrane of atrial gland DCVs.

Isolation and characterization of two homologous cDNA clones from Torpedo electromotor neurons.

Two homologous cDNA clones were isolated from a Torpedo california electric lobe lambda gt11 expression library using a polyclonal antiserum directed against proteins associated with synaptic vesicles. Northern blotting reveals an 8- to 9-kb transcript in the electric lobe and the spinal cord, but not in the brain or other non-neuronal tissues. Antibodies generated against a fusion protein synthesized in Escherichia coli reacted with a 85- to 90-kD species in the neurons of the electric lobe.

Modulation of ionic currents in Aplysia motor neuron B15 by serotonin, neuropeptides, and second messengers.

Both 5-HT and the 9 amino acid neuropeptide SCPb modulate 3 ionic currents in B15, enhancing a voltage-dependent inward sodium current, decreasing an outward potassium current and increasing an inward rectifying potassium current. In contrast, FMRFamide decreases a voltage-dependent inward sodium current and increases an outward potassium current. We have also investigated the roles of several second-messenger systems that may be mediating the effects of these modulators.

A bag cell neuron-specific antigen localizes to a subset of dense core vesicles in Aplysia californica.

The bag cell neurons of Aplysia govern egg-laying through the release of a number of bioactive peptides which are processed from a common precursor. Immunoelectron microscopic studies suggest that sorting at the trans-Golgi segregates peptides from the amino terminal and carboxy terminal of the precursor into distinct classes of dense-cored vesicles (DCVs). Here we identify a novel bag cell-specific antigen (4F6 antigen) using monoclonal antibodies (MAbs).

Two vesicle-associated membrane protein genes are differentially expressed in the rat central nervous system.

Vesicle-associated membrane protein (VAMP) 1 is a 120-amino acid protein which co-purifies with cholinergic synaptic vesicles from the marine ray Torpedo californica. We used the Torpedo gene to isolate two independent classes of VAMP cDNA clones from rat brain. Nucleotide sequence analysis of the cDNAs predicts proteins which are 84 and 75% homologous to Torpedo VAMP-1.

VAT-1: an abundant membrane protein from Torpedo cholinergic synaptic vesicles.

Expression screening was used to isolate cDNA clones encoding a synaptic vesicle membrane protein, VAT-1, which is specifically expressed in the electric lobe of marine rays. The predicted protein has a molecular weight of 41,572 daltons and contains several hydrophobic regions. An antibody raised against a fusion protein synthesized in E.

Multiple neuropeptides derived from a common precursor are differentially packaged and transported.

The ELH prohormone is proteolytically processed into at least nine peptides which govern egg-laying behavior in Aplysia. Quantitative immunocytochemistry demonstrates that peptides derived from the prohormone are packaged into distinct vesicle classes. Further experiments suggest the segregation occurs via a rapid initial proteolytic cleavage of the prohormone followed by sorting at the trans Golgi.

Processing of the egg-laying hormone (ELH) precursor in the bag cell neurons of Aplysia.

Egg laying in Aplysia is mediated by a battery of neuropeptides released from the bag cell neurons. Predominant intermediates in the proteolytic processing of the Aplysia egg-laying hormone neuropeptide precursor were characterized using biochemical and immunological techniques. Following removal of the signal peptide, a rapid cleavage at the tetrabasic sequence Arg-Arg-Lys-Arg separates the amino and carboxyl regions of the prohormone.

Synuclein: a neuron-specific protein localized to the nucleus and presynaptic nerve terminal.

We used an antiserum against purified cholinergic synaptic vesicles from Torpedo and expression screening to isolate a cDNA clone encoding synuclein, a 143 amino acid neuron-specific protein. A cDNA clone was also isolated from a rat brain cDNA library that encodes a highly homologous 140 amino acid protein. The amino terminal 100 amino acids of both proteins are comprised of an 11 amino acid repeating unit that contains a conserved core of 6 residues.

[Food chemistry and technology aspects of nickel-deficient nutrition in endogenously-induced allergic contact eczemas].

With reference to endogenous nickel contact dermatitis, the quantity of nickel resorbed is dependent on the individual capacity for absorption, the element species of nickel in the food matrix and the total nickel content in the food. As the content of nickel varies widely in individual foods, it is difficult to make any definite dietary recommendations. The wide fluctuation in the nickel content of vegetable food stuffs is due to variations in the composition of soil.

VAMP-1: a synaptic vesicle-associated integral membrane protein.

Several proteins are associated with, or are integral components of, the lipid bilayer that forms the delineating membrane of neuronal synaptic vesicles. To characterize these molecules, we used a polyclonal antiserum raised against purified cholinergic synaptic vesicles from Torpedo to screen a cDNA expression library constructed from mRNA of the electromotor nucleus. One clone encodes VAMP-1 (vesicle-associated membrane protein 1), a nervous-system-specific protein of 120 amino acids whose primary sequence can be divided into three domains: a proline-rich amino terminus, a highly charged internal region, and a hydrophobic carboxyl-terminal domain that is predicted to comprise a membrane anchor.

Egg-laying hormone, serotonin, and cyclic nucleotide modulation of ionic currents in the identified motoneuron B16 of Aplysia.

We have used the 2-electrode voltage-clamp technique to analyze the effects of the neuropeptide, egg-laying hormone (ELH), and the biogenic amine 5-HT on ionic currents in the buccal motoneuron B16 of Aplysia. When B16 is voltage-clamped near resting membrane potential, bath-applied ELH induces a prolonged inward shift in holding current. The ELH-induced inward current is not due to a decrease in the transient, the delayed, or the calcium-activated potassium currents.

Isolation and characterization of a Drosophila neuropeptide gene.

We have purified a 9 amino acid amidated neuropeptide, DPKQDFMRFamide, from whole adult D. melanogaster. This peptide exhibits sequence homology to the molluscan bioactive tetrapeptide FMRFamide and is a novel member of the FMRFamide peptide family.

Cellular and synaptic morphology of a feeding motor circuit in Aplysia californica.

The cellular and synaptic morphology of a component of the feeding motor circuit in Aplysia californica was examined with light and electron microscopic techniques. The circuit consists of a pair of inhibitory premotor interneurons, B4 and B5, as well as two motoneurons, B15 and B16, which innervate the accessory radula closer muscle. The neurons have wide, varicose arborizations in the buccal ganglion neuropil.

Aplysia californica neurons express microinjected neuropeptide genes.

Neuropeptide genes are expressed in specific subsets of large polyploid neurons in Aplysia californica. We have defined the transcription initiation sites of three of these neuropeptide genes (the R14, L11, and ELH genes) and determined the nucleotide sequence of the promoter regions. The genes contain the usual eucaryotic promoter signals as well as other structures of potential regulatory importance, including inverted and direct repeats.

Histidine-rich basic peptide: a cardioactive neuropeptide from Aplysia neurons R3-14.

We have previously demonstrated that neurons R3-14 of the Aplysia abdominal ganglia specifically express a gene encoding a 108-amino acid neuropeptide precursor. This precursor is postranslationally processed by cleavage of a signal sequence and two internal dibasic residues resulting in three peptides. The peptide products are colocalized in dense core granules throughout the R3-14 processes that innervate the efferent vein of the gill and the auricle.

Proteolytic processing of the Aplysia egg-laying hormone and R3-14 neuropeptide precursors.

A number of animal behaviors are influenced by the actions of neuropeptides that arise from the processing of complex protein precursors. In this report we investigate the proteolytic processing of neuropeptide precursors expressed in the Aplysia californica bag cells, which govern egg-laying, and neurons R3-14, which mediate aspects of cardiac output. Peptides were purified by fractionation on 2 high-pressure liquid chromatography systems followed by determination of amino acid compositions.

Peptidergic modulation of neuronal circuitry controlling feeding in Aplysia.

We examined the effects of 3 neuropeptides and the bioactive amine 5-HT on identified motoneurons (B15 and B16) and interneurons (B4, B5) involved in the control of feeding behavior in Aplysia californica. The application of egg-laying hormone (ELH), small cardioactive peptide b (SCPb), and 5-HT elicits distinct patterns of synaptically induced bursting in the neurons, while PheMetArgPheamide (FMRFamide) inhibits firing due to synaptic activity. Repetitive IPSPs recorded in B15 and B16 are induced by 5-HT and SCPb and inhibited by FMRFamide.

The Aplysia FMRFamide gene encodes sequences related to mammalian brain peptides.

We have characterized a precursor protein which gives rise to the neuropeptide Phe-Met-Arg-Phe-amide (FMRFamide) by determining the nucleotide sequence of a genomic and five cDNA clones. The 597-amino-acid protein contains 28 copies of the tetrapeptide FMRFamide, a single Phe-Leu-Arg-Phe-amide (FLRFamide), and other sequences flanked by paired basic residues, some of which have homologies to mammalian brain peptides. The data presented suggest the genes encoding pro-opiomelanocortin, pre-pro-enkephalin, and the hypothalamic releasing factor, cortico-releasing factor (CRF), arose from a common ancestral gene.