ELECTRON MICROSCOPIC STUDIES OF SILVER-STAINED PROTEINS IN NUCLEOLAR ORGANIZER REGIONS: LOCATION IN NUCLEOLI OF RAT SYMPATHETIC NEURONS DURING LIGHT AND DARK PERIODS

Ag-AS staining of nucleolar organizer regions was carried out on interphasic superior cervical ganglia neurons of rats sacrificed during light and dark periods. While the Ag-AS technique has mostly been used on monolayer cell lines or cell suspensions, the present study showed that in electron microscopy this technique is also applicable to small pieces of tissues. The finest pictures are obtained when (1) all solutions used for the staining procedure are at pH 4-5-4-7 and (2) the second step of the reaction involving ammoniacal silver and formalin developing solutions does not exceed 3 min. The results indicate that in the 2 time periods studied, a positive reaction took place exclusively in nucleolar fibrillar centres and in the fibrillar ribonucleoprotein (RNP) component (dense fibrillar component). The other nucleolar components, i.e. granular and vacuolar, were devoid of silver deposits. As previously shown in sympathetic neurons, the fibrillar centres of the nucleoli show a 10-fold increase in volume during the dark period. In this period, silver grains were located on both 'giant ' and small-sized fibrillar centres. The fibrillar RNP component seen either at the periphery of fibrillar centres or in the form of a well-delimited network showed the strongest reaction. The same distribution of silver grains was observed in the sympathetic neurons of rats sacrificed during the light period. Here again, silver accumulation occurred exclusively in the fibrillar centres and the fibrillar RNP component. The same difference in reactivity was observed as for the dark period, the fibrillar RNP component being the main site of the reaction