Granulosa cells (GCs) are somatic cells essential for establishing and maintaining bi-directional communication with the oocytes. This connection has a profound importance for the delivery of energy substrates, structural components and ions to the maturing oocyte through gap junctions. Cumulus cells, group of closely associated GCs, surround the oocyte and can diminished the effect of harmful environmental insults. Both GCs and oocytes prefer different energy substrates in their cellular metabolism: GCs are more glycolytic, whereas oocytes rely more on oxidative phosphorylation pathway. The interconnection of these cells is emphasized by the fact that GCs supply oocytes with intermediates produced in glycolysis. The number of GCs surrounding the oocyte and their age affect the energy status of oocytes. This review summarises available studies collaboration of cellular types in the ovarian follicle from the point of view of energy metabolism, signaling and protection of toxic insults. A deeper knowledge of the underlying mechanisms is crucial for better methods to prevent and treat infertility and to improve the technology of in vitro fertilization.

Abstract. Compared with advanced developmental stage embryos and somatic cells, fully grown mammalian oocytes contain specific nucleolus-like structures (NPB -nucleolus precursor bodies). It is commonly accepted that they serve as a store of material(s) from which typical nucleoli are gradually formed. Whilst nucleoli from somatic cells can be collected relatively easily for further biochemical analyses, a sufficient number of oocyte nucleoli is very difficult to obtain. We have found that isolated oocytes nucleoli fuse very efficiently when contact is established between them. Thus, well visible giant nucleoli can be obtained, relatively easily handled and then used for further biochemical analyses. With the use of colloidal gold staining, we estimated that a single fully grown mouse oocyte nucleolus contains approximately 1.6 ng of protein. We do believe that this approach will accelerate further research aiming at analyzing the composition of oocyte nucleoli in more detail. Key words: Fusion, Nucleolus, Oocyte, Protein content (J. Reprod. Dev. 58: [371][372][373][374][375][376] 2012) T he nucleolus is the most prominent structure in a cell nucleus [1]. In developmentally advanced embryos (i.e., blastocysts) and in somatic cells, it contains three basic components: fibrillar centers (FC), a dense fibrillar component (DFC) and a granular component (GC) [2]. Originally, it has been accepted that the nucleolus is exclusively the site of ribosomal RNAs (rRNAs) synthesis, processing and their assembly with ribosomal proteins, but now it is clear that nucleoli play an essential role(s) in many cellular processes [3]. Within the last few years, the composition of somatic cell nucleoli has been characterized, and it is now evident that somatic cell nucleoli contain thousands of proteins [4].Contrary to differentiated cells, fully grown mammalian oocytes and very early developing embryos prior to major embryonic genome activation do not contain typical tripartite nucleoli. Instead, socalled "nucleolus precursor bodies" (NPBs) are visible in germinal vesicles (GVs) and in pronuclei (PNs) of some mammalian species, i.e., rodents (mouse, rat), humans and pigs [5]. NPBs contain only a dense fibrillar mass. Contrary to somatic cell nucleoli, very little is known about the composition of NPBs. The only protein convincingly demonstrated to be present in them is nucleoplasmin 2 [6]. The function of NPBs in oocytes and embryos has been characterized, especially after the invention of so-called "enucleolation" [7]. This method represents the microsurgical removal of nucleoli from fully grown and growing oocytes as well as from one-cell stage embryos [8][9][10]. Thus, it has been convincingly demonstrated that NPBs are dispensable for maturation of fully grown oocytes. The enucleolation method also showed that nucleoli (NPBs) in zygotes are formed from the original oocyte nucleolar material [11]. Further detailed analysis demonstrated that the presence of nucleoli in pronuclei is absolutely essential for normal embryonic devel...

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The oocyte GV/GVs (germinal vesicle/germinal vesicles) and zygot PN/PNs (pronucleus/pronuclei) of some mammals contain clearly visible nucleoli which exhibit an atypical morphological structure. These nucleoli (NCLs) can be relatively easily manipulated, i.e. removed from GVs/PNs or eventually transferred into another oocyte/zygote. Thus, with the help of micromanipulation techniques it was possible to uncover the real function(s) they play in processes of oocyte maturation and early embryonic development. The purpose of our review is to describe briefly the micromanipulation techniques that can be used for oocyte/zygote nucleoli manipulation. Moreover, we present some examples of results that were obtained in nucleolus manipulation experiments.

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