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Distribution of FMRFamide-related peptides and co-localization with glutamate in Cupiennius salei, an invertebrate model system.

Cell and tissue research (2018-11-09)
Emily A Tarr, Brian M Fidler, Kyrstin E Gee, Carly M Anderson, Anna K Jager, Neil M Gallagher, Kaelyn P Carroll, Ruth Fabian-Fine
ABSTRACT

FMRFamide-related proteins have been described in both vertebrate and invertebrate nervous systems and have been suggested to play important roles in a variety of physiological processes. One proposed function is the modulation of signal transduction in mechanosensory neurons and their associated behavioral pathways in the Central American wandering spider Cupiennius salei; however, little is known about the distribution and abundance of FMRFamide-related proteins (FaRPs) within this invertebrate system. We employ immunohistochemistry, Hoechst nuclear stain and confocal microscopy of serial sections to detect, characterize and quantify FMRFamide-like immunoreactive neurons throughout all ganglia of the spider brain and along leg muscle. Within the different ganglia, between 3.4 and 12.6% of neurons showed immunolabeling. Among the immunoreactive cells, weakly and strongly labeled neurons could be distinguished. Between 71.4 and 81.7% of labeled neurons showed weak labeling, with 18.3 to 28.6% displaying strong labeling intensity. Among the weakly labeled neurons were characteristic motor neurons that have previously been shown to express ɣ-aminobutyric acid or glutamate. Ultrastructural investigations of neuromuscular junctions revealed mixed presynaptic vesicle populations including large electron-dense vesicles characteristic of neuropeptides. Double labeling for glutamate and FaRPs indicated that a subpopulation of neurons may co-express both neuroactive compounds. Our findings suggest that FaRPs are expressed throughout all ganglia and that different neurons have different expression levels. We conclude that FaRPs are likely utilized as neuromodulators in roughly 8% of neurons in the spider nervous system and that the main transmitter in a subpopulation of these neurons is likely glutamate.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Agarose, BioReagent, for molecular biology, low EEO
Sigma-Aldrich
Goat serum
Sigma-Aldrich
Bovine Serum Albumin, cold ethanol fraction, pH 5.2, ≥96%
Sigma-Aldrich
Mowiol® 4-88, Mw ~31,000
Sigma-Aldrich
Monoclonal Anti-Glutamate antibody produced in mouse, clone GLU-4, ascites fluid