Biochemical and immunocytological localization of the neuropeptides FMRFamide, SCPA, SCPB, to neurons involved in the regulation of feeding in Aplysia

Abstract

The localization of the neuropeptide FMRFamide in the buccal ganglia and buccal muscles of Aplysia was studied by immunocytology and high-pressure liquid chromatography (HPLC) combined with either a sensitive bioassay or 35S-methionine labeling. Immunocytology with an antiserum directed to FMRFamide stained a large number of fibers, varicosities, and neuronal somata. Two groups of stained neurons were of particular interest. One was the S cells, a group comprised of many small neurons, the majority of which were stained. HPLC of pooled labeled S cells confirmed that at least some of these neurons synthesize FMRFamide. The other group of stained neurons were in the ventral cluster, a group comprised of a small number of large neurons, many of which are motor neurons that innervate the buccal muscles involved in producing biting and swallowing movements. Several of the ventral neurons were previously shown to contain 2 other neuropeptides, the small cardioactive peptides SCPA and SCPB. These neurons are sufficiently large to permit HPLC analyses of the neuropeptides synthesized by individual neurons. This procedure confirmed that individual ventral neurons synthesized FMRFamide, or the SCPs, or all 3 peptides. The coexistence of FMRFamide and the SCPs in the same neuron was confirmed by simultaneous staining of sections from the buccal ganglia with a monoclonal antibody to the SCPs and an antiserum to FMRFamide. The coexistence of the 3 peptides in the same neuron was surprising in light of the observations that these peptides often have opposite biological activity. The ventral neurons are large and potentially identifiable as individuals. Thus, these neurons may be particularly useful for studying the physiological and behavioral roles of neuropeptides in generating complex behaviors.

Composite Modulatory Feedforward Loop Contributes to the Establishment of a Network State

“…Rostral view shows a cluster of small neurons that have been identified as radula mechanoafferent neurons (Miller et al 1994;Rosen et al 2000). Caudal view shows staining of large motor neurons including the two largest cells in the buccal ganglion, motoneurons B1 and B2, consistent with earlier findings (Lloyd et al 1987a(Lloyd et al , 1988. In addition, these immunohistochemical images indicate that the EN was strongly stained with SCP (see also Fig.…”

“…One source may be from the CNS because the SCP containing esophagealcluster neurons on the caudal surface ( Fig. 2A2), which include neurons B1 and B2, send their axons to the periphery through the EN (Lloyd et al 1987a(Lloyd et al , 1988. Note that the radula mechanoafferent neurons in the SCP-containing sensory neuron cluster on the rostral surface of buccal ganglion do not project to the EN.…”

Frequency-dependent release of peptide cotransmitters from identified cholinergic motor neurons in Aplysia.

“…One class of peptides that have been commonly found to coexist with other transmitters in Aplysia neurons are the small cardioactive peptides (SCPs) (6)(7)(8). The SCPs are represented by two peptides, SCPA and SCPB, which are 11 and 9 amino acids in length, respectively.…”

“…These peptides have similar sequences, are derived from a common precursor polypeptide (9)(10)(11), and have been shown to satisfy many of the criteria required of neurotransmitters (12). They are present in, and synthesized by, individual neurons (6)(7)(8) and are contained in membrane-bound dense-core vesicles in central neurons and in varicosities in the central nervous system, muscle, and gut (13,14). Immunoreactive fibers and varicosities are present at both central and peripheral sites where the SCPs have been shown to potently modulate the efficacy of synaptic transmission (12,15,16).…”

Pavlovian appetitive discriminative conditioning inAplysia californica

“…Moreover, Susswein and Byrne (1988) have characterized buccal cells that appear to serve as initiators of patterned activity consistent with that observed in the ingestion and rejection of food. Other work has succeeded in identifying several neuropeptides in the buccal cells (Cropper, Miller, et aI., 1990;Cropper, Tenenbaum, Kolks, Kupfermann, & Weiss, 1987;Lloyd, Frankfurt, Stevens, Kupfermann, & Weiss, 1987;Lloyd, Mahon, et aI., 1985) and in characterizing their modulatory effects on specific buccal neurons (Baux, Fossier, Trudeau, & Tauc, 1993;Sossin, Kirk, & Scheller, 1987).…”