Effect of environmental salinity and dopamine injections on key digestive enzymes in hepatopancreas of the euryhaline crab <i>Cyrtograpsus angulatus</i> (Decapoda: Brachyura: Varunidae)

Abstract

SUMMARY:We studied the occurrence and characteristics of lipase activity and the response of lipase and proteolytic activity to salinity and dopamine injections in hepatopancreas of the euryhaline crab (Cyrtograpsus angulatus). Lipase activity was maximal at pH 8.5; it exhibited Michaelis-Menten kinetics (apparent K m =0.019 mM), was higher at 37°C and appeared to be cold tolerant, being also high at 4°C. In 10 psu (hyper-regulation conditions), lipase and proteolytic activity were about 3 and 5 times higher, respectively, than in 35 psu (osmoconformation). In 40 psu (hypo-regulation), lipase activity was about three times higher than in 35 psu, while proteolytic activity was similar. Lipase activity was inhibited in vivo by 10 -4 M dopamine in 35 psu but not in 10 or 40 psu. Proteolytic activity was not affected by 10 -4 M dopamine. The differential responses of lipase and proteolytic activity to salinity and dopamine suggest the occurrence of distinct digestive adjustments and mechanisms of regulation upon osmoregulatory conditions. This study contributes to a better understanding of the complexity of the biochemical adaptations to salinity in euryhaline crabs. The fact that higher digestive enzyme activities could be associated with a differential digestive capacity potentially leading to enhanced availability of energy substrates is discussed.Keywords: Cyrtograpsus angulatus, euryhaline crabs, salinity, hepatopancreas, digestive enzymes, dopamine, Mar Chiquita lagoon. RESUMEN: EfEcto dE la salinidad ambiEntal y dE la inyEcción dE dopamina sobrE Enzimas digEstivas clavE En hEpatopáncrEas dEl cangrEjo EurihalinoCyrtograpsus angulatus (dEcapoda: brachyura: varunidaE). -Se estudió la existencia y características bioquímicas de actividad de lipasa y la respuesta a salinidad y a dopamina de actividad de lipasa y proteolítica total en el hepatopáncreas del cangrejo eurihalino Cyrtograpsus angulatus. La actividad de lipasa fue máxima a pH 8,5; mostró una cinética michaeliana (K m =0.019 mM); fue mayor a 37°C, y resistente a baja temperatura (4°C). En 10 psu (condición de hiperregulación), las actividades de lipasa y proteolítica fueron 3 y 5 veces mayores, respectivamente, que en 35 psu (condición de osmoconformación). En 40 psu (condición de hiporregulación), la actividad de lipasa fue de alrededor 3 veces mayor que en individuos aclimatados a 35 psu, mientras que la actividad proteolítica fue similar. La actividad de lipasa fue inhibida in vivo por dopamina 10 -4 M en 35 psu pero no fue afectada en 10 o 40 psu. Dopamina 10 -4 M no afectó la actividad proteolítica. Las respuestas de las actividades de lipasa y proteolítica a salinidad y a dopamina permiten sugerir la existencia de ajustes digestivos y mecanismos de regulación diferenciales en relación al estado osmorregulatorio. Los resultados constituyen un aporte relevante al conocimiento sobre las complejas adaptaciones a nivel bioquímico a la salinidad en cangrejos eurihalinos.

Amylase in the hepatopancreas of a euryhaline burrowingcrab: characteristics and modulation

“…Hemolymph (about 500 µL) was sampled from the infrabranchial sinus of 5-19 individuals by means of a syringe at the base of the cheliped and transferred to an iced centrifuge tube. Serum was separated by centrifugation at 10,000 × g for 30 s as described before (Michiels et al, 2013). Osmolality (mOsm kg -1 ) was measured in an aliquot of 50 µL of hemolymph and medium with a cryoscopic osmometer (Osmomat 030, Gonotec).…”

“…The modulation of digestive enzyme activities in the hepatopancreas (e.g., amylase) in response to different environmental salinity could lead to adjustments in the digestive capacity for specific substrates (starch/glycogen) and therefore in the availability of key metabolites (i.e. glucose) (Romano and Zeng, 2012;Michiels et al, 2013Michiels et al, , 2015aPinoni et al, 2015). However, studies at the molecular level, as we pointed out above, are lacking.…”

Biochemical characteristics and modulation by external and internal factors of aminopeptidase-N activity in the hepatopancreas of a euryhaline burrowing crab

“…modulation of digestive enzymes activities) upon changes in environmental conditions (Li et al 2008;Karasov et al 2011;del Valle et al 2004del Valle et al , 2006. However, in spite of their ecological importance studies in euryhaline estuarine crabs are still scarce and fragmentary (Asaro et al 2011;Michiels et al 2013;Pinoni et al 2013). Euryhaline crabs successfully occupying the intertidal area of coastal lagoons have to cope with abrupt and wide changes in environmental salinity.…”

“…Euryhaline crabs successfully occupying the intertidal area of coastal lagoons have to cope with abrupt and wide changes in environmental salinity. Biochemical adaptation to low and high salinity in such species appears to be a complex process involving the participation of enzymes and transport systems in branchial and extrabranchial tissues such as hepatopancreas and muscle (Jahn et al 2006;Pinoni and López Mañanes 2004Martins et al 2011;Athamena et al 2011;Michiels et al 2013;Pinoni et al 2013). However, some mechanisms of biochemical adaptation to low and high salinity, particularly from a metabolic perspective are still not fully understood.…”

“…absorption could lead to a greater availability of substrates (i.e. amino acid) for salinity acclimation (Li et al 2008;Romano and Zeng 2012;Michiels et al 2013). However, and strikingly, information about occurrence, biochemical characteristics or mechanisms of regulation of key proteolytic enzymes such as aminopeptidase-N in the hepatopancreas of euryhaline crabs is still lacking.…”