432 For transmission electron microscopy, specimens were fixed with 1.5% glutaraldehyde and 1.5% paraformaldehyde in 0.1 M sodium phosphate buffer, pH 7.3 for 3 h at room temperature and then postfixed for 2 h in 2% osmium tetroxide in 0.1 M sodium phosphate buffer. Samples were embedded in Araldite after dehydration in graded ethanol. To obtain a general overview of the samples semithin sections (1 μm) were cut with an Ultracut E (Leica), mounted on micro slides and stained with Methylenblue-Azur II for 1 min and viewed under an Axioplan 2 (Zeiss, Oberkochen, Germany) to which an Axiocam MRc 5 (Zeiss) was attached. Ultrathin sections were cut with an Ultracut E (Leica), mounted on copper grids, poststained with uranyl acetate and lead citrate and viewed under a LEO 900 (Zeiss). Results Description of the neurogenic PVZ of the anterior telencephalon A general overview of the telencephalon of larval X. laevis is given in Fig. 1a. To investigate the ultrastructural organization of the cell layers surrounding the lateral telencephalic ventricles we performed transmission electron microscopy. Figure 1b (b1) shows the anterior part of the lateral telencephalic ventricle, the PVZ and the posterior part of the granule cell layer of the OB. The transition between the anterior PVZ, adjacent to the OB, and the granule cell layer can be clearly identified (dotted line). Generally, the cells within the PVZ appear more electron dense than those in the adjacent cells layers. The neuronal stem-/progenitor cells within the anterior PVZ are arranged in four to five cell layers with irregular or oval nuclei that present, in some cases, small invaginations and prominent nucleoli (Fig. 1b, b2). Thereby, the cells contacting the ventricular lumen do not apparently differ from those within the other cell layers and are devoid of cilia or microvilli. The asterisk shows a neuronal stem-/progenitor cell in mitosis. Highly condensed DNA (chromosomes) is clearly visible showing that cells in the PVZ proliferate. The cell bodies of the granule cells have a more regular, roundish shape. The medial PVZ is made of about three to four cell layers and its cells basically resemble those of the anterior PVZ (Fig. 1b, b3). Some of them also have prominent nucleoli and they feature chromatin in different states of aggregation, probably representing pre- or post- mitotic stages of the cell cycle. As in the anterior part of the PVZ, also here the cells in direct contact with the lateral ventricle do not bear cilia or microvilli. The PVZ situated laterally to the ventricle appears thinner than in the anterior and medial part (Fig. 1b, b4). It is composed of more tightly packed cells arranged in at most two cell layers. Most cells Purinergic Signalling (2010) 6:429–445 have elongated nuclei and are devoid of cilia or microvilli. Figure 1b (b5 and b6) depicts higher magnification images of PVZ cells, two of them in mitosis, contacting the lumen of the lateral ventricle. The luminal parts of the cells are tightly packed with mitochondria. One cell exhibits a large cytoplasmic protrusion into the lumen of the ventricle; another cell exhibits a multivesicular protrusion expanding into the ventricle. This image shows also one of the rare cells extending a single cilium into the cerebrospinal fluid. To determine the proliferation rate of the PVZ cells and to further define the border between the neurogenic PVZ and the adjacent OB, we combined propidium iodide cell nuclei staining and a BrdU incorporation assay. In the adopted survey time window (see the “Materials and methods” section) the BrdU-positive cells, i.e., the proliferating cells, were confined to the PVZ. This helped to determine the approximate borders between the two brain regions (Fig. 2). Thereby, we found that the proliferation rate of PVZ cells is rather high. 42.54±6.65% (SEM; n=3 animals, 6 OBs/PVZs) of all cells surrounding the lateral ventricles were BrdU-positive. Nucleotide-induced increases of [Ca2+]i in cells of the anterior telencephalon Figure 3a (a1–a3) show a larva of X. laevis, a slice preparation of its telencephalon and a slice of the anterior part of the telencephalon stained with the fluorescent calcium-indicator dye Fura-2, respectively. To visualize nucleotide-induced [Ca2+]i signals in cells of the anterior telencephalon, we performed calcium imaging experiments. Figure 3b (b1) shows the same slice as in Fig. 3a (a3), with the red-shaded area covering the OB and the green shaded area covering the PVZ. Application of 5 μM 2MeSATP led to specific transient cellular response patterns in the vast majority of cells (Fig. 3b, b2). The response time courses of six individual cells of the OB and six cells of the PVZ are plotted in Fig. 3b (b3–b4). Figure 3b (b5) shows the mean (±SEM) intracellular Ca2+ transients of all 2MeSATP- responsive cells of this OB and PVZ. Application of ATP (50 μM) to the same slice preparation elicited similar, though smaller [Ca2+]i transients (Fig. 3b, b6). Similar results were obtained in more than 20 slices of the anterior telencephalon tested for their responsiveness to 2MeSATP and ATP. These results strongly suggest that both, OB cells and PVZ cells, are endowed with purinergic receptors. To exclude purinergic receptor desensitization, interstimulus intervals of at least 5 min were chosen in all of the experiments. Fig. 3c shows that ATP-induced [Ca2+]i increases remain stable using this experimental protocol. To confirm that the nucleotide-induced responses were truly mediated by purinergic receptors, we tested the effects of two acknowledged P2 receptor antagonists. In both brain regions, the OB and the PVZ, 2MeSATP- and ATP-induced