Ganias et al,: Degeneration of postovulartory follicles in Sordino pikhordus 



137 



Figure 7 



Consecutive stages of oocyte atresia in the Iberian sardine Sardina pilchardiis from very new 

 (A), to intermediate (B), to very late (C, D). T=theca layer; G = granulosa layer; arrow indicates 

 the aggregation of very late atretic oocytes in the central area of an ovarian lamella. Scale bar: 

 0.1 mm. 



all stages, atretic follicles constitute enclosed cellular 

 structures (Fig. 7), whereas POFs always maintain an 

 opening towards the ovarian lumen (Fig. 3). In addition, 

 as previously reported, late atretic oocytes separate 

 from the epithelium of the lamellae (Fig. 7D), whereas 

 POFs remain on the epithelium until full resorption 

 (Fig. 3). 



Sardine POFs shrank exponentially with time, reduc- 

 ing to almost half their size daily. Therefore, after the 

 exponential relationship of POF resorption with time 

 is estimated, ages may be estimated by applying the 

 data for the cross-sectional areas of POFs and times of 

 capture to the model. The measurement of POF cross- 

 sectional areas is not very time consuming and could 

 be merged into the routine of histological analysis for 

 DEPM analyses. Finally, the attribution of ages may 

 be finely tuned by inspecting histomorphological char- 

 acteristics, especially from specimens that lie outside 

 the fitting curve. 



Before applying the above aging procedure to fish 

 populations, attention should be drawn to other fac- 

 tors that may affect POF cross-sectional area, such 

 as the laboratory treatment of ovaries and the envi- 

 ronmental conditions at sampling, e.g., water tempera- 



ture. In our study, the use of different preservation 

 media (AFA solution and formalin) did not appear 

 to affect the size of the POFs. On the other hand, 

 the embedding material significantly affected the 

 follicle's size, for POFs in resin were almost double 

 those in paraffin. Resin is known to maintain cellu- 

 lar organization in tissues slightly affected, whereas 

 paraffin causes significant shrinkage (Casotti, 2001; 

 Dorph-Petersen et al., 2001). Besides size, the mor- 

 phological characteristics of the follicles also differed 

 between the two materials. Although the whole POF 

 structure remained compact in resin, follicular folds 

 in paraffin had shrunk and were detached from the 

 thecal layer, even in young POFs. These differences, 

 in combination to the thinner slices that are achieved 

 with resin, make resin a much better material for de- 

 tailed histological observations. However, for accuracy 

 in the staging of POFs, both resin and paraffin seem 

 to provide similar results. 



Water temperature may affect the rate of POF resorp- 

 tion in teleosts both in the field (Ganias et al., 2003; 

 Roumillat and Brouwer, 2004) and laboratory studies 

 (Fitzhugh and Hettler, 1995). However, this effect has 

 never been precisely quantified and thus temperature 



