Process Of Synapse Elimination Research Articles

The role of gonadal hormones in neuromuscular synapse elimination in rats. I. Androgen delays the loss of multiple innervation in the levator ani muscle

The normal period of synapse elimination in the androgen-sensitive levator ani (LA) muscle occurs between 2 and 4 weeks after birth, well after the period of synapse elimination for most other rat muscles. To evaluate whether gonadal androgen might be involved in the delayed development of single innervation in the LA, we compared the time course of synapse elimination in LA muscles that lacked endogenous gonadal androgen or were exposed to exogenous androgen. Tetranitroblue tetrazolium was used to stain neuromuscular connections. Our results suggest that both endogenous and exogenous androgen delay the normal process of synapse elimination. Removing endogenous androgen resulted in lower levels of multiple innervation in the LA, suggesting that androgen may normally influence synapse elimination. Moreover, androgen treatment prevented much of the normal loss of multiple innervation in the LA. Androgen treatment during the normal period of synapse elimination also increased the diameter of LA muscle fibers, enhanced the development of preterminal branching, and increased the number of junctional sites on some LA fibers. Because androgen did not appear to induce the formation of new synapses through sprouting, we conclude that androgen maintained multiple innervation in the LA by preventing the normal ontogenetic process of synapse elimination.

On the role of the 200-kDa neurofilament protein at the developing neuromuscular junction

To examine whether the 200-kDa neurofilament protein (200K NFP) is involved in mechanically stabilizing axons, we studied the developmental appearance of immunoreactivity to nonphosphorylated and phosphorylated 200K NFP at the neuromuscular junction. Polyinnervated rat muscle fibers become singly innervated during the first 3 weeks of postnatal life through the process of synapse elimination. If production or post-translational modification of the 200K NFP is actively involved in imparting mechanical stability on neuromuscular synapses, then the selective presence of this protein in only one of several axons at each developing end plate region might make that one axon selectively resistant to elimination. The remaining axons would then be eliminated. Immunoreactivity to the 200K NFP is present on Gestational Day 14 and can be seen in more than one preterminal axon in the end plate region of a muscle fiber during the period of synapse elimination. These results suggest that the 200K NFP is present and phosphorylated early in development and, although the 200K NFP may increase the mechanical stability of axons, this increased stability does not determine the final outcome of synapse elimination.

Synapse elimination occurs late in the hormone-sensitive levator ani muscle of the rat.

Using tetranitroblue tetrazolium (TNBT) to stain neuromuscular synapses, we compared the development of the adult pattern of innervation in two fast-twitch muscles in the rat: the androgen-sensitive levator ani (LA) and the extensor digitorum longus (EDL), which is not thought to be androgen sensitive. We found that about 18% of adult LA muscle fibers, but only about 2% of adult EDL fibers, are multiply innervated. Moreover, synapse elimination occurs substantially later in the LA compared with the EDL. At 2 weeks after birth, the EDL is already predominantly singly innervated, whereas the LA is still predominantly multiply innervated. The apparent delay in the normal time course of synapse elimination in the LA corresponds to a similar delay in other aspects of neuromuscular development (the time course of appearance of axonal retraction bulbs, the growth of fibers, and the development of adult motor terminal morphology). Finally, motor terminals change during synapse elimination from morphologies resembling growth cones to the adult form of neuromuscular synapses. Because the period of synapse elimination is significantly different for muscles that differ in their androgen sensitivity, hormonal sensitivity may represent an important property of motoneurons or muscle fibers influencing the normal time course of neuromuscular synapse elimination in rats. Thus, androgen might regulate the normal ontogenetic process of synapse elimination.

Competition favouring inactive over active motor neurons during synapse elimination.

During normal postnatal maturation, mammalian muscles undergo an orderly process of synapse elimination, whereby each muscle fibre loses all but one of the multiple inputs with which it is endowed at birth. Experimental perturbations that increase or decrease the overall activity of nerve and/or muscle cause a corresponding increase or decrease in the overall rate of neuromuscular synapse elimination. On other grounds it has been suggested that competition among motor neurons is important in determining which synapses survive and which are eliminated. Would a difference in activity among the terminals at the same endplate affect the outcome of the competition and not just its rate? We investigated this issue by blocking activity for four days in a small fraction of the motor neurons innervating the neonatal rabbit soleus muscle. Twitch tensions of motor units were subsequently measured for both the active and inactive populations of neurons to assess whether the inactive neurons had lost fewer or more synapses than is normal. We found that inactive motor neurons have a significant advantage compared to active counterparts in control experiments, a finding opposite to that expected if the neuromuscular junction operated by classical 'Hebbian' rules of competition.

Early postnatal development of the monkey visual system. II. Elimination of retinogeniculate synapses

Profiles of retinal terminals, and of their synaptic and non-synaptic contacts, were measured in electron micrographs from magnocellular and parvocellular laminae of the dorsal geniculate nucleus (LGNd) in newborn, 1-, 4-, 8- and 17-week-old rhesus monkeys. Morphologic criteria, i.e., the presence of pale mitochondria and large round vesicles, were used to identify the profile of retinal origin. Size-frequency histograms were stereologically reconstructed and used to calculate the density of retinal boutons and synaptic and non-synaptic plaques. The density values were adjusted for laminar growth to yield estimates of total numbers of these elements. Numerical estimates indicate bouton proliferation during the first week, followed by substantial reductions in bouton number accompanied by profound decreases in synapse number and cumulative synaptic area. In magnocellular layers, the reduction in synapse number is more pronounced after the eighth week, whereas the decrements in both features in the parvocellular laminae occur before this time. This synapse elimination process may be due entirely to retinal bouton retraction in parvocellular layers, but involves additional retinal synapse loss in the magnocellular segment. The parvocellular division shows a further size contraction of the remaining synapses. Immature synapses predominate in the LGNd throughout the 4-month period, and no quantitative evidence for direct transformation of immature to mature contacts is obtained. Non-synaptic junctions are stable in number but of increasing size in magnocellular layers, whereas substantial increases in number and area are found in parvocellular laminae. The preceding modifications in synaptic organization of the monkey LGNd occurring during the initial postnatal period may provide morphologic bases for the physiological and behavioral changes observed in this species during the same interval. Our data underscore the conclusion that synaptic reorganization occurs over a prolonged period, probably extending beyond 4 months, and involving the process of synapse elimination.

Postnatal development of the inferior olivary complex in the rat. II. Topographic organization of the immature olivocerebellar projection.

The state of organization of the olivocerebellar projection in newborn and 5-day-old rats has been analyzed by autoradiography of anterogradely transported 3H-leucine, as well as by retrograde transport of horseradish peroxidase. The efferent axons of the inferior olivary neurons are already present and already highly organized in the cerebellum of newborn rats. Most of the autoradiographic labelling subsequent to the injection of 3H-leucine into the inferior olive is seen in the subcortical medullary zone. Labelled axons only partially invade the gray matter, where they reach the zone occupied by randomly distributed Purkinje cells. At this immature stage, olivocerebellar projections are already entirely crossed and distributed according to a pattern which is similar to the adult. At the fifth postnatal day olivocerebellar projections have moved from the medullary zone toward the interface between the molecular and the granular layers where Purkinje cells have arranged in a monolayer. Evidence for translocation of climbing fibers from their perisomatic to their peridendritic position is already distinct in these young cerebella. Combination of anterograde and retrograde fiber system tracing experiments discloses the following crossed topography of olivocerebellar projections: The caudal half of the medial accessory olive projects mainly to the vermis of the posterior lobe, whereas its rostral half projects to the flocculus, paraflocculus, and the intermediate cortex. The principal olive, ventral and dorsal lamellae, supplies climbing fiber inputs to the hemispheric cortex. The caudal half of the dorsal accessory olive projects to the lateral portion of the vermis of the anterior lobe, whereas neurons in its rostral half send their axons toward the intermediate cortex. This topographic arrangement is, therefore, similar to that reported for adult mammals. The present results, alone or when compared with those obtained during other studies on the synaptogenesis between climbing fibers and Purkinje cells, allow the following conclusions: The climbing fibers enter the cerebellar cortex before Purkinje cells have reached the developmental phase compatible with synaptogenesis. They wait in the medullary white matter until appropriate maturation of their cellular targets. Olivocerebellar topography is roughly similar in newborn, 5-day -old, and adult rats. Synaptogenesis between climbing fibers and Purkinje cells, which is known not to start before the second postnatal day, is not necessary for the establishment of the topographic organization of the olivocerebellar projection.

Reduction of multiaxonal innervation at the neuromuscular junction of the rat during development.

1. A study was made of the functional and structural changes that occur during the decrease in multiaxonal innervation of neonate rat muscle fibres 2-14 days after birth. 2. In day 8 to day 14 animals there was a constant daily loss in the average number of functionally transmitting axons/muscle fibre measured electrophysiologically. An investigation of synaptic transmission during this period revealed that the loss of functional contact from the supernumerary axons was not preceded by any sign of failing terminal conduction or a gradual decrease in transmission efficacy but rather appeared to occur abruptly. 3. Neonate end-plates showing signs of abnormal ultrastructure were observed during the period of synapse elimination. Some axon terminals had a high cytoplasmic density and condensation of synaptic vesicles. Signs of Schwann cell encroachment into the synaptic cleft were readily found and large areas of post-junctional membrane apposed only by Schwann cell were evident. 4. It is suggested that the mechanics of the process of synapse elimination in neonates is similar to that occurring during degeneration in the denervated adult. Transmission failure occurs abruptly at the supernumerary endings and they are disposed of by the Schwann cell.