An improved method for long-term measuring of hemolymph fluctuations of non-essential amino acids, GABA and histamine from freely moving crayfish

The microdialysis method was adapted to obtain long-term hemolymph dialysates from the pericardial cavity of freely moving Procambarus clarkii crayfish, to measure fluctuations of non-essential amino acids, GABA and histamine by high-performance liquid chromatography using off-line fluorometric deri...

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Bibliographic Details
Published in:Journal of neuroscience methods 2006-05, Vol.153 (1), p.1-7
Main Authors: Cebada, Jorge, Alvarado-Álvarez, Ramón, Becerra, Elizabeth, Neri-Bazán, Leticia, Rocha, Luisa, García, Ubaldo
Format: Article
Language:English
Subjects:
Amino Acids - analysis
Amino Acids, Essential - analysis
Animals
Astacoidea - physiology
Behavior, Animal - physiology
Cells, Cultured
Circadian fluctuations
Crayfish hemolymph
GABA
gamma-Aminobutyric Acid - analysis
Hemolymph - metabolism
Histamine
Histamine - analysis
Locomotion - physiology
Microdialysis
Microdialysis - instrumentation
Microdialysis - methods
Non-essential amino acids
Procambarus clarkii
Reproducibility of Results
Sensitivity and Specificity
Time Factors
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Summary:The microdialysis method was adapted to obtain long-term hemolymph dialysates from the pericardial cavity of freely moving Procambarus clarkii crayfish, to measure fluctuations of non-essential amino acids, GABA and histamine by high-performance liquid chromatography using off-line fluorometric derivatization. Asp, Ala, Tau, GABA and histamine (HA) reached its maximal concentrations at the daybreak, whereas glutamate (Glu), Gln and Gly peaked at the end of the light period. The minimum and maximal detected amounts for each substance along the 24 h cycle were (in μM): 20–300 Asp, 100–200 Glu, 400–700 Gln, 400–600 Gly, 100–200 Tau, 150–300 Ala, 2–10 GABA and 25–250 HA. Cocktails containing the relative concentration of each amino acid, GABA and histamine resulted in a hyperpolarization that reduced the spontaneous firing of cultured peptidergic X organ neurons. Glu, GABA and histamine evoked a long-lasting hyperpolarization that suppressed the spontaneous firing, whereas Asp, Gly and Tau evoked a depolarization accompanied with neuronal firing. Finally, neither Ala nor Gln modified the resting membrane potential.
ISSN:0165-0270
1872-678X
DOI:10.1016/j.jneumeth.2005.05.025