The P2X 7 receptor (P2X 7 R) is a purinoceptor expressed predominantly by cells of immune origin, including microglial cells. P2X 7 R has a role in the release of biologically active proinflammatory cytokines such as IL-1β, IL-6 and TNFα. Here we demonstrate that when incubated with lipopolysaccharide (LPS), glial cells cultured from brain of P2X 7 R −/− mice produce less IL-1β compared to glial cells from brains of wild-type mice. This is not the case for TNFα and IL-6. Our results indicate a selective effect of the P2X7R gene deletion on release of IL-1β release but not of IL-6 and TNFα. In addition, we confirm that only microglial cells produce IL-1β, and this release is dependent on P2X 7 R and ABC1 transporter. Because IL-1β is a key regulator of the brain cytokine network and P2X 7 R is an absolute requirement for IL-1β release, we further investigated whether response of brain cytokines to LPS in vivo was altered in P2X 7 R −/− mice compared to wild-type mice. IL-1β and TNFα mRNAs were less elevated in the brain of P2X 7 R −/− than in the brain of wild-type mice in response to systemic LPS. These results show that P2X7R plays a key role in the brain cytokine response to immune stimuli, which certainly applies also to cytokinedependent alterations in brain functions including sickness behavior.

No abstract

In order to determine at which follicular size goat oocytes were capable of resuming and completing meiosis, we evaluated their ability to mature in vitro and measured their maturation promoting factor (MPF) activity by histone H1 kinase assay. The results indicated that goat oocyte meiotic competence developed progressively in follicles ranging from 0.5 to 3 mm; the oocytes acquired the ability to resume meiosis in follicles of 0.5-0.8 mm, to reach metaphase I (MI) in follicles of 1-1.8 mm, and to reach metaphase II (MII) in follicles larger than 3 mm. The presence of MPF activity was first observed in oocytes arrested at early prometaphase I and reached a maximum level in oocytes blocked in metaphase I (MI). In the second part of this study, RNA synthesis and nucleolar changes were analyzed during the growth period. The acquisition of meiotic competence was accompanied by nucleolar compaction and a dramatic decrease in RNA synthesis. Changes in protein patterns were also analyzed, but only slight differences were observed among oocytes from the different classes.

Microinjection of bacterially expressed human cdc25A protein into Xenopus prophase oocytes provokes the activation of p34cdc2 kinase and the tyrosine dephosphorylation of p34cdc2 in the presence or absence of protein synthesis. The level of p34cdc2 kinase activity then drops in parallel with the degradation of cyclin B2 and finally increases again to stabilize at a high level. Cdc25 microinjection induces the assembly of a metaphase I spindle which is abnormally located in the deep cytoplasm. Moreover, oocytes arrest at the metaphase I stage and do not reach metaphase II even 10 h after cdc25 microinjection. The extended metaphase I period observed in cdc25-injected oocytes results from an equilibrium between degradation of cyclins and synthesis of new cyclins. This is in contrast with progesterone-stimulated oocytes where cyclin degradation is turned off when oocytes enter metaphase II. During metaphase I, the reactivation of MPF activity can be disrupted in two different ways: 1) cycloheximide, an inhibitor of protein synthesis, by preventing the synthesis of new cyclins, provokes the disappearance of MPF kinase activity and the reformation of a nucleus; 2) when the cAMP level is increased during the metaphase I period in cdc25-injected oocytes, MPF kinase activity drops following a rephosphorylation of tyrosine 15 of p34cdc2, while the cyclin turn-over remains unaffected. Moreover, increasing the cAMP level in prophase oocytes totally prevents the action of cdc25. Our results indicate that in Xenopus oocytes, the PKA pathway negatively regulates the activation of MPF and the activity of p34cdc2/cyclin B complex through tyrosine phosphorylation of p34cdc2 during metaphase I.