Distribution of serotonin-immunoreactivity in juvenile Aplysia

Goldstein, RS; Kistler, Henry B.; Steinbusch, H.W.M.; Schwartz, James H.

doi:10.1016/0306-4522(84)90043-5

Cited by 93 publications

(47 citation statements)

References 29 publications

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“…For a second animal maintained in continuous darkness, the diurnal rhythm of 5-HT sulfate also remained intact. Given these results, and as serotonergic fibers innervate the eye in marine mollusks (Goldstein et al 1984;Takahashi et al 1989), we sought to understand whether the source of the 5-HT sulfate was from the eye itself. We examined whether 5-HT sulfate was produced in the eye by incubating one eye with 5-HT and the second with only PS from the same animal.…”

Section: -Htp Injections Elevate Circulating 5-ht Sulfate In Freely mentioning

confidence: 99%

Systemic serotonin sulfate in opisthobranch mollusks

Stuart¹,

Ebaugh²,

Copes³

et al. 2004

Journal of Neurochemistry

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Serotonin (5-hydroxytryptamine, 5-HT) is a ubiquitous modulatory neurotransmitter with roles as a neurohormone and neurotransmitter. However, few studies have been performed characterizing this molecule and its related metabolites in circulating fluids. Here, we demonstrate native 5-HT sulfate, but much lower levels of 5-HT, in hemolymph of the marine mollusk Pleurobranchaea californica. The metabolite 5-HT sulfate forms from 5-HT uptake and metabolism in central ganglia of Aplysia californica and in the visceral nerve and eye of Pleurobranchaea, but not in hemolymph itself. In addition, 5-hydroxyindole acetic acid (5-HIAA), while not detected in hemolymph, forms in higher quantities than does 5-HT sulfate in the eye and visceral nerve, and c-glu-5-HT is also observed in this area but never in hemolymph. As systemic 5-HT sulfate appears not to originate from the optic region or from systemic 5-HT, 5-HT sulfate likely derives from the nervous system. Circulating 5-HT sulfate is at least 10-fold higher during the light portion of a 12 : 12-h light/dark cycle than during the dark portion (p < 0.0007), but there is no obvious trend for free systemic tryptophan (Trp) (p > 0.3) in Pleurobranchaea. 5-HT in mollusks is associated with general arousal state; thus, diurnal systemic changes in a 5-HT catabolite may reflect a regulatory role for indole catabolism in behavioral rhythms. Keywords: capillary electrophoresis, hemolymph, marine mollusk, serotonin, sulfation. Serotonin (5-hydroxytryptamine, 5-HT) is a biogenic amine responsible for a variety of tasks in the nervous system. For instance, 5-HT is implicated in learning and memory in the central nervous system (CNS) and emesis and peristaltic reflex in the enteric nervous system (ENS) (Bartsch et al. 2000;Buhot et al. 2000;Endo et al. 2000). Several organs serve as targets for 5-HT, but the fate of 5-HT in target tissues, let alone the physiological effects of the resulting metabolites, has not been fully ascertained. Knowledge of the products of transformed 5-HT, as well as their distribution patterns, will assist in illuminating the processes leading to impaired neurotransmission and eventually aid in designing compounds that could potentially inhibit disease in neural and surrounding tissues.Marine mollusks present an excellent model for the study of the nervous system because of their large cell somata and well-characterized neural networks. In gastropod mollusks, 5-HT is involved in many neurophysiological processes, including sensitization and facilitation of withdrawal reflexes (Hawkins et al. 1993), feeding behavior (Gillette and Davis 1977) and locomotion (McPherson and Blankenship 1991). However, 5-HT-related metabolites have received considerably less attention.Our laboratory recently demonstrated that 5-HT metabolism in Aplysia californica involves sulfation and c-glutamylation with trace amounts of monoamine oxidase activity present in the form of 5-hydroxyindole acetic acid (5-HIAA) (Stuart et al. 2003). In particular, 5-HT sulfate is present ...

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Section: -Htp Injections Elevate Circulating 5-ht Sulfate In Freely mentioning

confidence: 99%

Systemic serotonin sulfate in opisthobranch mollusks

Stuart¹,

Ebaugh²,

Copes³

et al. 2004

Journal of Neurochemistry

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“…Immunocytochemistry was done by using the methodology described by Goldstein et al (7). In brief, the tissue was fixed in a phosphate (0.1 M)-buffered solution of sucrose (30%) and paraformaldehyde (4%), and 30-,um frozen sections were taken.…”

Section: Methodsmentioning

confidence: 99%

Evidence for parallel actions of a molluscan neuropeptide and serotonin in mediating arousal in Aplysia.

Lloyd¹,

Kupfermann²,

Weiss³

1984

Proc. Natl. Acad. Sci. U.S.A.

119

116

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The neuropeptide designated SCPB (small cardioactive peptide B), the sequence of which has recently been determined, was found in the accessory radula closer muscle, a muscle involved in biting movements. The ganglia and nerves that innervate the accessory radula closer muscle also contain SCPB. At nanomolar concentrations, it enhances the contractions of the muscle. The effect of SCPB on the muscle resembles the effect of an identified serotonergic neuron that previously was shown to mediate behavioral effects that reflect a food arousal state in Aplysia. Like serotonin, SCPB enhances contractions by a postsynaptic action, which appears to involve an increase in cAMP levels in the muscle. Our findings suggest that parallel peptidergic and serotonergic pathways may mediate similar aspects of arousal in Aplysia.Gastropod molluscs contain a number of neuropeptides that have been identified on the basis of their powerful cardioactive actions (1). One of these peptides, termed small cardioactive peptide B (SCPB) was recently purified from the neural tissue of Aplysia (2) and its sequence has been determined (Met-Asn-Tyr-Leu-Ala-Phe-Pro-Arg-Met-NH2). The present results describe our first functional study utilizing synthetic SCPB.We have found that SCPB is present in particularly high levels in the buccal ganglion of Aplysia, the ganglion that controls the muscles that effect biting and swallowing (3,4). Therefore, we have examined whether SCPB might be involved in the regulation of these muscles. The accessory radula closer (ARC) muscle was chosen for study because this muscle has been examined physiologically and behaviorally (4,5). The ARC muscle has been used as a model system for studying the peripheral aspects of a behavioral arousal state elicited by food. Previous research has shown that food-induced arousal in Aplysia is associated with a serotonergic input to the ARC muscle and that serotonin enhances the force of contraction of this muscle. We now report that SCPB enhances the contractile response of this muscle to motor neuron stimulation in a manner similar to that of serotonin.We also determined that SCPB is present in neural elements in the ARC muscle and in the nerve that innervates it. These findings suggest that a peptidergic system using SCPB may function in parallel to the serotonergic system to mediate aspects of food-induced arousal. METHODSTo determine the levels of SCPB in muscle, ganglia, and nerves, the tissues were homogenized and purified with HPLC, and the levels of SCPB were measured by bioassay. Fresh or frozen tissue was placed in at least a 20-fold excess of 0.1 M acetic acid, heated to 100°C for 10 min, and homogenized. The supernatant of a 10-min, 10,000 x g centrifugation of the homogenate was lyophilized, taken up in a measured volume of 0.1 M acetic acid, and subjected to reversephase HPLC on a Zorbax C8 column (DuPont) using a DuPont 850 pump and gradient controller. The column was developed with an acetonitrile gradient from 25% to 50% at 1%/min at a flow rate of 1 ml/m...

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“…Immunocytology of cultured neurons was done using techniques previously described for whole mounts and sections of Aplysia tissue (7,14). In brief, individual cells or groups of cells growing in culture were fixed for 3 hr with 4% paraformaldehyde in 0.1 M phosphate Abbreviations: RP-HPLC, reversed-phase high-performance liquid chromatography; SCP, small cardioactive peptide.…”

Section: Methodsmentioning

confidence: 99%

Release of neuropeptides during intracellular stimulation of single identified Aplysia neurons in culture.

Lloyd¹,

Schacher²,

Kupfermann³

et al. 1986

Proc. Natl. Acad. Sci. U.S.A.

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An important criterion for classifying a substance as a neurotransmitter is that it is released in an activity-dependent fashion. We have utilized cell culture of individual neurons of Aplysia to demonstrate the release of the neuropeptides SCPA and SCPB (small cardioactive peptides A and B). Neurons B1 and B2 were isolated from the buccal ganglion of Aplysia and maintained in cell culture. The cells grew new processes, which were immunoreactive to antibodies for the neuropeptide SCPB. These processes contained SCPA and SCPB that were detectable by bioassay on snail heart. The cells synthesized the SCPs from radiolabeled precursors and transported the peptides to their neurites. Single cells released SCPs in a calcium-dependent fashion upon intracellular electrical stimulation. Taken together, these results provide critical evidence that SCPs are neurotransmitters. The results also indicate that the cell culture of individual identified neurons can be used to investigate the release of peptides.The small cardioactive peptides (SCPs) are a class of low molecular weight neuropeptides that have been shown to have potent modulatory actions both in the central nervous system and on peripheral tissues in Aplysia (1-4). To date, two SCPs have been purified and sequences: SCPA (Ala-ArgPro-Gly-Tyr-Leu-Ala-Phe-Pro-Arg-Met-NH2; ref. 5) and SCPB (Met-Asn-Tyr-Leu-Ala-Phe-Pro-Arg-Met-NH2; ref. 6). The carboxyl-terminal amide and seven amino acids are conserved between the two peptides. The two SCPs are present in the same neurons (7), an observation consistent with molecular genetic analyses which indicate that a precursor protein contains the sequences of both peptides (8).The SCPs have been shown to satisfy many of the criteria required of chemical mediators. These peptides are present in and synthesized by individual neurons (7) and are contained in membrane-bound dense core vesicles (9). Immunocytology indicates that the SCPs are not ubiquitously distributed within the nervous system but rather are confined to a select population of neurons. Immunoreactive fibers and varicosities are found at both central and peripheral sites where the SCPs have been shown to potently modulate the efficacy of synaptic transmission (1, 3). The SCPs, however, have not been shown to be released from neurons in a calciumdependent manner. Although there is not a consensus on the precise criteria that must be satisfied to prove that a substance is a neurotransmitter (see ref. 10 for a discussion of these criteria), one criterion that is generally considered to be crucial is that the substance is released during neural activity.An additional corollary of this criterion is that the release should be dependent on extracellular calcium concentrations.Because transmitters, especially peptides, are released in small quantities, the demonstration of neurotransmitter release is often facilitated by using radiolabels. Labeling of traditional transmitters is further facilitated by the fact that their precursors are rapidly taken up at synaptic te...

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Distribution of serotonin-immunoreactivity in juvenile Aplysia

Cited by 93 publications

References 29 publications

Systemic serotonin sulfate in opisthobranch mollusks

Systemic serotonin sulfate in opisthobranch mollusks

Evidence for parallel actions of a molluscan neuropeptide and serotonin in mediating arousal in Aplysia.

Release of neuropeptides during intracellular stimulation of single identified Aplysia neurons in culture.

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