Ectonucleotidase activities in Sertoli cells from immature rats

• Published in: Brazilian journal of medical and biological research 2001-10, Vol.34 (10), p.1247-1256
• Main Authors: Casali, E A, da Silva, T R, Gelain, D P, Kaiser, G R, Battastini, A M, Sarkis, J J, Bernard, E A
• Format: Article
• Language: English
• Subjects: 5'-Nucleotidase - metabolism; Adenine Nucleotides - metabolism; Adenosine Deaminase - metabolism; Adenosine Diphosphate - metabolism; Adenosine Monophosphate - metabolism; Adenosine Triphosphate - metabolism; Animals; ATP diphosphohydrolase; BIOLOGY; Chromatography, High Pressure Liquid - methods; ecto-5'-nucleotidase; ectoadenosine deaminase; Hydrolysis; Male; MEDICINE, RESEARCH & EXPERIMENTAL; purine nucleotides; Rats; Rats, Wistar; Sertoli cells; Sertoli Cells - enzymology
• ISSN: 0100-879X; 1414-431X; 0100-879X; 0034-7310
• DOI: 10.1590/S0100-879X2001001000003

Sertoli cells have been shown to be targets for extracellular purines such as ATP and adenosine. These purines evoke responses in Sertoli cells through two subtypes of purinoreceptors, P2Y2 and P A1. The signals to purinoreceptors are usually terminated by the action of ectonucleotidases. To demonstrate these enzymatic activities, we cultured rat Sertoli cells for four days and then used them for different assays. ATP, ADP and AMP hydrolysis was estimated by measuring the Pi released using a colorimetric method. Adenosine deaminase activity (EC 3.5.4.4) was determined by HPLC. The cells were not disrupted after 40 min of incubation and the enzymatic activities were considered to be ectocellularly localized. ATP and ADP hydrolysis was markedly increased by the addition of divalent cations to the reaction medium. A competition plot demonstrated that only one enzymatic site is responsible for the hydrolysis of ATP and ADP. This result indicates that the enzyme that acts on the degradation of tri- and diphosphate nucleosides on the surface of Sertoli cells is a true ATP diphosphohydrolase (EC 3.6.1.5) (specific activities of 113 +/- 6 and 21 +/- 2 nmol Pi mg(-1) min(-1) for ATP and ADP, respectively). The ecto-5'-nucleotidase (EC 3.1.3.5) and ectoadenosine deaminase activities (specific activities of 32 +/- 2 nmol Pi mg(-1) min(-1) for AMP and 1.52 +/- 0.13 nmol adenosine mg(-1) min(-1), respectively) were shown to be able to terminate the effects of purines and may be relevant for the physiological control of extracellular levels of nucleotides and nucleosides inside the seminiferous tubules.