Comparative Biochemistry and Physiology Part B: Biochemistry and Molecular Biology
Branchial carbonic anhydrase (CA) of gills of Chasmagnathus granulata (Crustacea Decapoda)
Introduction
Carbonic anhydrase (CA) is a ubiquitous enzyme in vertebrates in which it has been intensively studied, particularly in mammals. On the other hand, the information about occurrence, characteristics and physiological role in invertebrates is still fragmentary.
In decapod crustaceans, CA has been involved in all of the major gill functions such as CO2 excretion, acid-balance and ionoregulation (Henry, 1988a, Henry, 1988b, Botcher and Siebers, 1993, Botcher et al., 1995). The pattern of distribution of CA activity among gills pairs appears to vary depending on habitat and osmoregulatory behavior of the animal (Henry, 1988a, Henry, 1988b, Henry and Cameron, 1982a, Botcher and Siebers, 1993). In euryhaline species such as Callinectes sapidus (Henry and Cameron, 1982a), Carcinus maenas (Botcher et al., 1990a, Botcher et al., 1990b) Eriocheir sinensis (Olsowoski et al., 1995) and in crayfish Pacifistacus leniusculus (Wheatly and Henry, 1987) CA activity was described to be strongly dependent on environmental salinity. Upon acclimation of individuals to reduced salinity CA activity enhanced in posterior gills whereas appeared not to be affected in anterior gills (Botcher and Siebers, 1993). Furthermore, after an abrupt change to dilute media CA activity has been described to increase in posterior gills seven of C. sapidus (Henry and Cameron, 1982b). However, to our knowledge, only in P. lenisculus the response of CA activity to an abrupt salinity change in each individual gill pair has been studied (Henry and Wheatly, 1988).
Several biochemical, physiological and pharmacological studies support the existence of two pools of CA in euryhaline crabs gills — a cytosolic CA which has been suggested to have an important role in posterior gills ion uptake by providing H+ and HCO3− for apical ion transport systems and a membrane-associated CA probably involved in CO2 excretion (Henry, 1988a, Henry, 1988b, Botcher et al., 1990a, Botcher et al., 1990b, Botcher et al., 1991, Botcher et al., 1995, Botcher and Siebers, 1993, Onken and Riestenpatt, 1998). The existence and predominance of these two forms of CA appear to vary depending on species. In aquatic and semiterrestrial euryhaline crustacean gills the distribution of the cytoplasmic pool of CA appears to be a function of the euryhalinity of the species and the ion transporting capability of the tissue whereas the presence of membrane-associated CA activity would be related to a high metabolic rate (CO2 production) and an active lifestyle (Henry, 1991).
In C. maenas the predominant membrane-bound CA has been shown to exhibit biochemical and pharmacological features similar with mammalian membrane-bound CA IV (Botcher et al., 1994).
Chasmagnathus granulata (Grapsidae) is a euryhaline semiterrestrial crab which is found from southern Brazil to Patagonia (Argentina) (Boschi, 1964, Botto and Irigoyen, 1969). In Mar Chiquita coastal lagoon (Buenos Aires Province, Argentina) it is one of the dominant crabs in the outer parts where it is exposed to highly and abruptly variable environmental salinity ranging from 4 to 36‰ (Anger et al., 1994, Spivak et al., 1994).
Although C. granulata has been identified as an hyper–hypo osmotic regulator and it is known to support a wide range of environmental salinity (Mañe-Garzón et al., 1974, Gnazzo et al., 1978, Luquet et al., 1992) its branchial ionoregulatory mechanisms at the biochemical level are poorly understood.
The aim of this work was to determine the occurrence, localization and response to environmental salinity of CA activity in all of the gills of C. granulata from Mar Chiquita lagoon.
Section snippets
Chemicals
4-2(hydroxyethyl)-1-piperazinethane sulphonic acid (Hepes) was from Boehringer (Manheim, Germany); sucrose was from Fluka (Germany); ethylenediamine tetraacetic acid (EDTA), Tris–(hydroxymethylamino-methane) (Tris); ethylene glicol N,N′,N′-tetracetic acid (EGTA), acetazolamide and bovine serum albumin were from Sigma (St. Louis, MO, USA). All chemicals were of analytical grade. All solutions were prepared in glass-distilled water.
Animal collection and maintenance
Crabs were caught from a single area of Mar Chiquita lagoon. Only
Localization and distribution of CA activity in gills of C. granulata
CA specific activity was determined in gills 1+2+3 and each of the gills pairs 4–8 of C. granulata acclimated for 3 weeks to either 35 or 10‰ salinity, salinities to which this crab is usually exposed in its natural environment. In 35‰ salinity CA specific activity was homogeneously distributed among individual gills 4–8 and pooled gills 1+2+3. On the other hand, in individuals acclimated to 10‰ salinity highest CA specific activity (about 90–100 μm H+ min−1 per mg protein) was found in
Discussion
Results herein described show the occurrence of CA activity sensitive to salinity in each gills of the intertidal crab C. granulata from Mar Chiquita lagoon (Fig. 1, Fig. 2). The more uniform level of CA activity among gills of C. granulata acclimated to 35‰ salinity when hemolymph osmolality and Na+, K+ and Cl− concentrations are about the same of external medium (Table 1) is in accordance with that described in the euryhaline crab C. sapidus exposed to conditions in which it behaves as
Acknowledgements
We wish to thank Mabel Garcı́a for her technical assistance during the writing of this paper. This work was in part supported by a grant from the University of Mar del Plata, Argentina.
References (35)
A rapid and sensitive method for the quantitation of microgran quantities of protein-dye binding
Anal. Biochem.
(1976)- et al.
Branchial Na+–K+ATPase and osmoregulation in the purple shore crab (Hemigrapsus nudus)
Comp. Biochem. Physiol.
(1996) Non-cytoplasmic carbonic anhydrases
TIBS
(1988)- et al.
pH-dependent association of carbonic anhydrase (CA) with gastric light microsomal membranes isolated from bovine abomasum. Partial characterization of membrane-associated activity
Comp. Biochem. Physiol.
(1993) - et al.
Transepithelial potential differences and sodium fluxes in isolated perfused gills of the mangrove crab (Ucides cordatus)
Comp. Biochem. Physiol.
(1998) - et al.
NaCl absorption across split gill lamellae of hyperregulating crabs: transport mechanisms and their regulation
Comp. Biochem. Physiol.
(1998) - et al.
Enhancement of carbonic anhydrase activity by erythrocyte membranes
Arch. Biochim. Biophys.
(1989) - et al.
Carbonic anhydrase inhibitors. Part 52. Metal complexes of heterocyclic sulfonamides — a new class of strong topical intraocular pressure-lowering agents in rabbit
Eur. J. Med. Chem.
(1998) - et al.
Hatching rhythms and dispersion of decapod crustacean larvae in a brackish coastal lagoon in Argentina
Helgolander Meeresunters
(1994) Sodium–potassium activated adenosine triphosphatase and cation transport
Los crustáceos decápodos brachyura del litoral bonaerense (R. Argentina)
Boln. Inst. Biol. Mar. Mar del Plata.
Biochemistry, localization and physiology of carbonic anhydrase in the gills of euryhaline crabs
J. Exp. Zool.
Carbonic anhydrase in branchial tissues of osmoregulating shore crab (Carcinus maenas)
J. Exp. Zool.
Localization of carbonic anhydrase in the gills of (Carcinus maenas)
Comp. Biochem. Physiol.
Physiological role of branchial carbonic anhydrase in the shore crab (Carcinus maenas)
Mar. Biol.
Membrane-associated carbonic anhydrase from the crab gill: purification, characterization and comparison with mammalian CAs
Arch. Biochem. Biophys.
Carbonic anhydrase, a respiratory enzyme in the gills of the shore crab (Carcinus maenas)
Helgolander Meeresunters
Cited by (30)
Trypsin and N-aminopeptidase (APN) activities in the hepatopancreas of an intertidal euryhaline crab: Biochemical characteristics and differential modulation by histamine and salinity
2017, Comparative Biochemistry and Physiology -Part A : Molecular and Integrative PhysiologyMolecular cloning and sequence analysis of two carbonic anhydrase in the swimming crab Portunus trituberculatus and its expression in response to salinity and pH stress
2016, GeneCitation Excerpt :Na+/K+-ATPase is combined with biological membrane; CA is divided into two groups, one is named cytoplasmic carbonic anhydrase (CAc) and the other one is named glycosyl-phosphatidylinositol-linked carbonic anhydrase (CAg). CAc exists in cytoplasm and CAg is combined with biological membrane (Jillette et al., 2011;López Mañanes et al., 2000). Except for the difference in distribution, both of the molecular structure of the two kinds of CA contain four primary components: the zinc binding site (Christianson and Alexander, 1989), the threonine-199 loop (Merz, 1990), the substrate association pocket (Krebs et al., 1993) and the proton shuttling mechanism (Tu and Silverman, 1989).
Carbonic anhydrase induction in euryhaline crustaceans is rate-limited at the post-transcriptional level
2014, Comparative Biochemistry and Physiology - A Molecular and Integrative PhysiologyCitation Excerpt :In general, significant CA induction takes several days to occur. For example, in the euryhaline marine crab Chasmagnathus granulata, low salinity-stimulated CA activity does not occur until 3–8 days post-transfer (Mañanes et al., 2000). A 2004 study by Palacios et al. found no difference in CA activity in the gills of the white shrimp Litopenaeus vannamei within 3 or 24 h of low salinity transfer; however later studies would show that significant induction occurs after 7 days (Roy et al., 2007).
Between-habitat comparison of digestive enzymes activities and energy reserves in the SW Atlantic euryhaline burrowing crab Neohelice granulata
2011, Comparative Biochemistry and Physiology - A Molecular and Integrative PhysiologyCitation Excerpt :Both chelae were cut off and carapaces were removed. The hepatopancreas, chela muscle, anterior (1–5) and posterior (6–8) gills (López Mañanes et al., 2000) were immediately excised and weighed. Wet mass was measured to the nearest 0.01 g.
Na<sup>+</sup> ATPase activities in chela muscle of the euryhaline crab Neohelice granulata: Differential response to environmental salinity
2009, Journal of Experimental Marine Biology and EcologyBiochemical and physiological adaptations in the estuarine crab Neohelice granulata during salinity acclimation
2008, Comparative Biochemistry and Physiology - A Molecular and Integrative PhysiologyCitation Excerpt :As for Na+/K+-ATPase, carbonic anhydrase activity of all gills (anterior and posterior gills) is also dependent on the environmental salinity. The pattern of carbonic anhydrase distribution among gills changes from a more uniform distribution when crabs are osmo- and iono-conforming their body fluids to a highest enzyme activity in posterior gills (6–8) when crabs are hyper-regulating these fluids (Mañanes et al., 2000; Genovese et al., 2005a). Therefore, a differential involvement of the carbonic anhydrase present in anterior and posterior gills in the ionoregulatory mechanisms is suggested to occur in N. granulata.