Release of a Neurosecretory Hormone as Peptide by Electrical Stimulation of Crab Pericardial Organs

1. Saline which had bathed an isolated crab pericardial organ was chromatographed on a column of Sephadex G-25. The fractions were divided and assayed for cardioexcitor activity and for ninhydrin colour following hydrolysis. 2. Fluid from pericardial organs which had been stimulated electrically to give a maximum propagated compound action potential showed both cardio-excitor activity and ninhydrin colour. These were always in the same fractions, which corresponded to the volume for elution of small peptides. Fluid from unstimulated preparations gave negative assays. 3. The cardio-excitor activity of fluid from stimulated pericardial organs and of distilled-water homogenates of pericardial organs behaved identically in Sephadex G-25 chromatography. 4. No evidence could be obtained of the cardio-excitor peptide binding to a protein. 5. This work provides evidence that neural stimulation of crab pericardial organs results in release to the bathing fluid of cardio-excitor hormone as free peptide.

Do the Monoamines in Crab pericardial Organs Play a Role in Peptide Neurosecretion?

1. Pericardial organs of Libima emargmata and L. dubia were isolated, the nerves from the thoracic ganglion were stimulated electrically, and propagated electrical activity was recorded. Fluid which bathed the neurohaemal organ was assayed for cardio-excitor effects on the isolated crab heart, when necessary, made tachyphylactic to drugs. 2. Neither 10-6M 5-hydroxytryptamine (5-HT), 10-5M dopamine (DA), nor 10-5M UML applied directly to the pericardial organ evoke release, nor do DA or UML alter release in response to stimulation. 3. The rate of cardio-excitor release is normal in pericardial organs taken from reserpinized animals. Histochemical examination of the pericardial organs by the formaldehyde-induced fluorescence method confirmed depletion of the monoamines. 4. These experiments, taken together, provide strong evidence against a role of monoamines in the release of cardio-excitor hormone from this neurosecretory structure.

Fluorescence Localization of Monoamines in Crab Neurosecretory Structures

1. The pericardial organs and anterior ramifications (both neurohaemal structures) of six species of crabs have been examined as whole mounts by the histochemical method for monoamines based on formaldehyde-induced fluorescence. 2. A small number of specifically fluorescing axons (not more than six green and six yellow) innervate the pericardial organ; one of the green-fluorescent and one yellow-fluorescent axon branches and also innervates the anterior ramification. 3. All of the fluorescing axons enter via segmental nerves 1, 2 and 3 from the ventral ganglion. 4. One large, brilliant green-fluorescing axon, and the small green-fluorescing axon which branches to the AR, have been traced in Carcinus to cell bodies in the circumoesophageal connective ganglion. These cells may give rise to the entire population of green-fluorescing axons and terminals in the neurohaemal organs. 5. Each axon, throughout its course in the pericardial organ, supplies a dense array of varicosities (blebs) at surfaces which are directly exposed to the haemolymph. The anterior ramifications are also supplied with blebs. 6. Lack of fluorescence in controls not exposed to paraformaldehyde, reversible quenching of fluorescence by treatment with sodium borohydride, and depletion of the fluorescence by reserpinization of the crabs, all confirm that the fluorescence is specific and represents the intracellular localization of monoamines. 7. With the aid of data available elsewhere we conclude that there are distributed, in parallel with peptide-secreting axons and terminals in the pericardial organs and anterior ramifications, a group of dopamine-containing and a group of 5-hydroxytryptamine-containing axons and terminals.