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

Abstract

Strikingly, in spite of its physiological importance, information about occurrence, biochemical characteristics and mechanisms of regulation of aminopeptidase-N (APN) in the hepatopancreas of intertidal euryhaline crabs is still lacking. In this work, we determined the occurrence, biochemical characteristics, response to environmental salinity and dopamine of APN in the hepatopancreas of the euryhaline crab Neohelice granulata (Dana 1851) from the open mudflat of Mar Chiquita coastal lagoon (Buenos Aires province, Argentina). APN activity was maximal at pH and temperature range of 7.6-9.0 and 37-45 °C, respectively. APN activity exhibited Michaelis-Menten kinetics (apparent Km = 0.19 ± 0.10 mM) (pH 7.6, 37 °C) and appeared to be sensitive to bestatin (I 50 = 15 mM) and EDTA (I 50 = 9 mM). In crabs acclimated to 10 psu (hyper-regulation conditions) and 37 psu (hypo-regulation conditions), APN activity was about 45 and 160% higher, respectively, than in 35 psu (osmoconformation). APN activity in the hepatopancreas was stimulated in vitro (about 137%) by 10(-4) M dopamine. Higher dopamine concentrations produced a similar extent of increase. The responses of APN activity to salinity and dopamine in vitro suggest the role of APN in digestive adjustments upon hyper and hypo-regulatory conditions and its modulation via direct mechanisms on hepatopancreas by dopamine.

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

“…The differences in the number of amylase forms present in the hepatopancreas of crustaceans have been associated with variations in environmental and/or physiological factors (Borowsky, 1984;Le Moullac et al, 1997;Perera et al, 2008aPerera et al, , 2008bCastro et al, 2012). N. granulata behaves as a hyper/hyporegulator since it exhibits hemolymph osmolarity values higher and lower than those of the corresponding external medium upon acclimation to 6-10 psu and 37-60 psu, respectively, while it osmoconforms at 35 psu (Asaro et al, 2011;Pinoni et al, 2013;Michiels et al, 2015a;Pinoni et al, 2015; this work, Table). The additional 30 kDa species found by zymogram analysis in the hepatopancreas from crabs acclimated to low salinity (10 psu) (hyperregulation conditions) (Figure 1b) suggests that mechanisms of modulation at the molecular level occur, which could lead to the higher total amylase activity detected by biochemical analysis.…”

The euryhaline crabUca tangerishowed metabolic differences to sex and environmental salinity

“…type of reserves, storage organs involved) are species-specific and depend on the type of osmotic challenge (Romano & Zeng, 2012; Pinoni et al ., 2013; Romano et al ., 2014). Hyper and hypo-osmoregulation can require different mechanisms which lead to differential adjustments in carbohydrates, protein and lipid metabolism (Michiels et al ., 2013, 2015; Pinoni et al ., 2013, 2015; Romano et al ., 2014). The possible occurrence of differential metabolic adjustments related to sex is poorly studied.…”

“…Decapod crustaceans use carbohydrates, lipids and proteins as principal energy molecules but the extent of use and the relative importance of the different metabolic sources involved vary depending on species and type of challenge (Pinoni et al ., 2013; Jimenez & Kinsey, 2015; Wang et al ., 2016). In males of various species of decapod crustaceans, adaptation to environmental salinity is a complex process involving adjustments at the biochemical level in several tissues and organs and a metabolic reorganization (Michiels et al ., 2013, 2015; Pinoni et al ., 2013; Chen et al ., 2015). Modifications in various haemolymph metabolites have been reported (Romano et al ., 2012, 2014; Larsen et al ., 2014; Prymaczok et al ., 2016).…”

Digestive flexibility in a euryhaline crab from a SW Atlantic coastal lagoon: alkaline phosphatase activity sensitive to salinity in the hepatopancreas

“…In some euryhaline species, the modulation of different digestive enzymes activities in the hepatopancreas in response to low salinity suggests the occurrence of digestive and metabolic adjustments at the biochemical level which could lead to a greater availability of substrates (i.e. aminoacids) for salinity acclimation (Li et al ., 2008; Asaro et al ., 2011; Romano & Zeng, 2012; Michiels et al ., 2013, 2015a, b; Pinoni et al ., 2013; Wang et al ., 2014). Alkaline phosphatases (AP) (EC 3.1.3.1) are membrane-bound glycoproteins that are widely found in animals which hydrolyse phosphate from a variety of molecules at a range of optimal pH above 7.0 (Ali et al ., 2013; Buchet et al ., 2013; Linder et al ., 2013).…”

“…In Mar Chiquita coastal lagoon (Argentina), N. granulata is one of the dominant crabs inhabiting areas with abrupt, frequent and highly variable changes in salinity (Spivak et al ., 1994; Iribarne et al ., 1997; Bortolus and Iribarne 1999; Fanjul et al ., 2008; Luppi et al ., 2013). Previous work from our lab shows that complex and integrative responses occur upon acclimation to low salinity (López Mañanes et al ., 2000; Schleich et al ., 2001; Pinoni et al ., 2005, 2013; Pinoni & López Mañanes, 2009; Asaro et al ., 2011; Michiels et al ., 2015a). In chela muscle of N. granulata levamisole-insensitive AP activity is differentially modulated by low salinity suggesting its role in adjustments underlying hyper-regulation (Pinoni et al ., 2005).…”