Ultrastructural characterization of synaptosomes from neonatal and adult rats with special reference to monoamines

Abstract

The crude mitochondria1 fraction P2 and subfractions of it were prepared from the brain stem, hemispheres, hypothalamus and striatum of adult rats and from the hemispheres and brain stem or diencephalon-mesencephalon and pons-medulla oblongata of 1-day-old rats. Samples were fixed in glutaraldehyde-osmium, KMnO, or by Tranzer's triple fixation method (aldehydes-chromate-dichromate-osmium) and examined by electron microscopy. With all these fixation techniques 80-95% of processes in the adult samples were found to be mature synaptosomes. After incubation with either 10-100 pg/ml of 5-hydroxydopamine or a-methylnoradrenaline, and after permanganate or triple fixation, monoamine synaptosomes, containing small granular vesicles were detected.They were found in all the brain regions studied and in the fraction P2 and subfractions C and D but only a t a frequency of less than 1%. In samples from l-dayold rats using the different fixatives, synaptosomes comprised 10-30% of all processes. Nerve endings were a t very different stages of maturation in sharp contrast to the rather uniform type of adult synaptosomes. They varied from processes still without any substructures to mature synaptosomes containing synaptic junctions. Monoamine synaptosomes containing small granular vesicles were detected after amine incubation and KMnO, or triple fixation. Their frequency was lower than that in the adult. Most common were very immature terminals containing smooth endoplasmic reticulum and a very few small granular vesicles, indicating a very early appearance of the capacity to form small granular vesicles under experimental conditions. The immature terminals had all the characteristics of the growth cone and might partly have originated from growth cones a t the tips of growing axons and dendrites. Less frequent were more mature synaptosomes containing a greater number of small granular vesicles and no longer any smooth endoplasmic reticulum. The crude microsomal fraction P3 was also examined in the adult brain regions and the brain stem of 1-day-olds.This fraction contained a small number of small agranular vesicles originating from cell bodies and nonterminal axons; their number was smaller in 1-day-old than in adult rats. The present results are in agreement with the high proportion of immature monoamine terminals in newborn rat brain detected earlier by fluorescence histochemistry.Earlier studies have demonstrated that density, contrasting to the rather uniform synaptosomes in a low proportion can be iso-adult type of synaptosomes (Spence and lated from the brain of newborn rats.

Ultrastructure of synaptosomes from fetal rat brain

“…Contrary to our previous results in the 1-day-old rat (Kanerva et al 1976(Kanerva et al , 1977 small DCV's could not be demonstrated after incubation in either 5-OHDA or a-methyl-NA. This reflects the immaturity of the monoamine-storing vesicle pool at this stage of development.…”

“…We have recently confirmed and extended the results of Jones and Revell (1970) and Gonatas et al (1971) in the 1-day-old rat , Kanerva et al 1976, 1977. The purpose of this further study of the ontogenesis of synaptosomes was partly to analyze the fine structure of fetal rat synaptosomes.…”

“…It has been suggested that the tubular reticulum which is believed to store the non-granular pool of monoamines (Eranko 1972, Tranzer 1972) is labile and the homogenization procedure could thus liberate their amine content during the processing of synaptosomes. We have previously demonstrated small granular vesicles in the 1-day-old rat although their number was small (Kanerva et al 1976(Kanerva et al , 1977. Varicose nerve terminals have been demonstrated in the fetal rat brain by the formaldehyde-induced fluorescence (FIF) technique (Seiger and Olson 1973).…”

“…They might be part of the smooth endoplasmic reticulum and/or represent developing or degenerating synaptic vesicles . Synaptosomes packed with so-called small granular vesicles (SGV) typical of monoaminergic nerve endings (Hokfelt et al 1970, Kanerva et al 1976, 1977 have not been observed in the fetal rat.…”

Developmental changes in the activity of phosphatidylethanolamine N-methyltransferases in rat brain

“…1). The subcellular origin of the preparation termed 'synaptosomal' in that study is also uncertain, since it has been shown that, although one can prepare from neonatal animals (by means of centrifugations in discontinuous sucrose density gradients), a fraction whose density is equivalent to that of synaptosomes from the adult animals, these preparations are not at all similar to synaptosomes prepared from adult brains (as assessed by electron microscopy and marker enzyme assays) (Jones & Revell, 1970;Gontas et al, 1971;Kanerva et al, 1977); their subcellular origin is unclear (Jones & Revell, 1970;Gontas et al, 1971;Kanerva et al, 1977).…”