Fine structure of synaptogenesis in the vertebrate central nervous system.

This article reviews studies of the formation of synaptic junctions in the vertebrate central nervous system. It is focused on electron microscopic investigations of synaptogenesis, although insights from other disciplines are interwoven where appropriate, as are findings from developing peripheral and invertebrate nervous systems. The first part of the review is concerned with the morphological maturation of synapses as described from both qualitative and quantitative perspectives. Next, epigenetic influences on synaptogenesis are examined, and later in the article the concept of epigenesis is integrated with that of hierarchy. It is suggested that the formation of synaptic junctions may take place as an ordered progression of epigenetically modulated events wherein each level of cellular affinity becomes subordinate to the one that follows. The ultimate determination of whether a synapse is maintained, modified or dissolved would be made by the changing molecular fabric of its junctional membranes. In closing, a hypothetical model of synaptogenesis is proposed, and an hierarchial order of events is associated with a speculative synaptogenic sequence. Key elements of this hypothesis are 1) epigenetic factors that facilitate generally appropriate interactions between neurites; 2) independent expression of surface specializations that contain sufficient information for establishing threshold recognition between interacting neurites; 3) exchange of molecular information that biases the course of subsequent junctional differentiation and ultimately results in 4) the stabilization of synaptic junctions into functional connectivity patterns.

Ectopic dendritogenesis and associated synapse formation in swainsonine-induced neuronal storage disease.

Ectopic dendrite growth and new synapse formation are known to occur on select kinds of neurons in a wide variety of neuronal storage diseases. As these changes in connectivity occur just proximal to the axonal initial segment, it has been hypothesized that they underlie the generation of abnormal neuronal function in these diseases. We have studied certain aspects of this phenomenon through the use of a plant-derived indolizadine alkaloid, swainsonine, which specifically inhibits the lysosomal hydrolase, alpha-mannosidase. These studies fully document the close morphological similarity between swainsonine-induced and inherited feline alpha-mannosidosis. This includes the presence of clear and floccule-filled storage vacuoles, as seen with routine EM, and axon hillock neurite growth on select cell types, as seen with Golgi staining. The latter was found only on cortical pyramidal neurons and multipolar cells of amygdala, and these same cell types are known to be involved in ectopic neuritogenesis in other storage diseases. Combined Golgi-electron-microscopic studies demonstrated the presence of normal-appearing synapses on these aberrant neuritic processes and also unusual, membranous inclusions specifically within the neurite-bearing pyramidal cells. The latter may be indicative of unique metabolic changes in these neurons and is consistent with the hypothesis that storage of gangliosides or other glycolipids underlies the recapitulation of dendritic growth features in these diseases. Experimental manipulation of the disease process using the swainsonine model indicated that induction of cortical pyramidal neuron neurite growth could be influenced by both age of onset and intensity of intraneuronal storage. Although Golgi studies clearly demonstrated neuritic sprouting in animals with disease onset as late as at 1 year, cortical pyramidal cells of older, adult animals appeared to undergo significant storage without a similar induction of neurite growth. These studies support the view that induced neuritogenesis in neuronal storage disease is associated with changes in metabolism, specifically within the neurite-bearing cells, that this change possibly involves gangliosides, and that the neuritogenic response may be limited to pre-adult stages of brain maturation.

Regulation of motor axon sprouting.

Our studies on the amphibian and mammalian motor systems suggest that sprouting of intact motoneurons and synapse formation can be regulated by three mechanisms: peripheral, central, and transneuronal. Peripheral mechanisms provide the means of a direct mode of interaction between the periphery of the nerve cell and the target, to determine the extent of target innervation. The central mechanism enables target muscles to signal the cell bodies of their innervating motoneurons to regulate axonal growth and synapse formation, and thus again determine the extent of their innervation. The transneuronal mechanism provides a vehicle by which the pattern of innervation of a muscle can be altered by nerve cells that do not themselves innervate the muscle, but are an integral part of the entire system.

Elimination of action potentials blocks the structural development of retinogeniculate synapses.

Although the influence of electrical activity on neural development has been studied extensively, experiments have only recently focused on the role of activity in the development of the mammalian central nervous system (CNS). Using tetrodotoxin (TTX) to abolish sodium-mediated action potentials, studies on the visual system show that impulse activity is essential both for the normal development of neuronal size and responsivity in the lateral geniculate nucleus (LGN), and for the eye-specific segregation of geniculo-cortical axons. There have been no anatomical studies to investigate the influence of action potentials on CNS synaptic development. We report here the first direct evidence that elimination of action potentials in the mammalian CNS blocks the growth of developing axon terminals and the formation of normal adult synaptic patterns. Our results show that when TTX is used to eliminate retinal ganglion-cell action potentials in the cat from birth to 8 weeks, the connections made by ganglion cell axons with LGN neurones, retinogeniculate synapses, remain almost identical morphologically to those in the newborn kitten.

Transplantation of fetal lateral geniculate nucleus to the occipital cortex: connectivity with host's area 17.

The developing lateral geniculate complex was excised from fetal albino rats at 18 days of gestation and implanted into the occipital cortex of host animals at 5 days of postnatal age. Groups of host animals were sacrificed at 10, 20 and 30 days following this procedure. The transplant tissue of selected animals was stereotaxically lesioned 2 days prior to scheduled sacrifice and their brains subjected to either Fink-Heimer or electron microscopic analysis of the distribution and density of degenerating efferents from the transplant. The remaining animals were analysed by means of Bodian, Golgi-Cox or electron microscopic techniques. Transplanted neurons displayed typical dendritic branching patterns of geniculate relay neurons by 20 days following implantation. Intrinsic neurons, characterized by a small ovoid soma and two main stem dendrites, only became evident in transplant tissue by 30 days and were much reduced in number. Synapses developed by 10 days and rapidly increased in number by 20 and 30 days. Most complexes were simple axo-dendritic, asymmetric junctions. Multiple serial and reciprocal complexes, as well as the characteristic glomerular complex, failed to appear. Analysis of Bodian stained material revealed a dense network of fibers coursing about the transplant. Distinct bundles of these fibers were observed extending from the medial edge of the transplant into area 17 by 20 days following implantation. A Fink-Heimer analysis of animals whose transplants were stereotaxically lesioned revealed degeneration in Layers II-VI of the primary visual cortex but the majority of these fibers terminated within the lateral two-thirds of Layer IV. Few degenerated fibers could be found in the underlying white matter indicating that efferents from the transplant found their way to their "correct" target zone by growing through a complex neuropil which provided minimal physical substrates to guide such growth. Most of the contacts formed by these fibers were simple junctions along the shafts of dendrites with a wide range in diameter. It is concluded that the nearby host visual neurons, which are the correct target cells for the afferents arising in the transplant, induced a directed growth of these fibers.

Changes in MEPP and EPP amplitude distributions in the mouse diaphragm during synapse formation and degeneration.

Miniature end-plate potential (MEPP) and end-plate potential (EPP) amplitude histograms were examined in the mouse diaphragm during degeneration, deterioration, re-innervation and neonatal development. MEPPs and EPPs were recorded with conventional electrophysiological techniques. Control MEPP amplitude distributions from mice 21-30 days old showed two classes of MEPPs. The larger class composed 80-90% of the MEPPs and formed a bell-shaped distribution (bell-MEPPs). The smaller class (skew-MEPPs) formed a skewed distribution with a peak 1/7 to 1/15 that of bell-MEPPs. Usually, MEPP amplitude distributions did not change during the course of nerve degeneration or during deterioration in the bath. MEPP amplitude distributions from newly re-innervated fibers were composed mainly of skew-MEPPs. At later stages of re-innervation the relative numbers of skew-MEPPs decreased. Many fibers from neonatal mice (2-3 days old) also showed mainly skew-MEPPs. Rise time vs amplitude plots were constructed from neonatal and re-innervating preparations. The skew-MEPP time-to-peak measurements fell on or below the regression line calculated from the time-to-peak data of the bell-MEPPs. This indicates that the skew-MEPPs originated from the same site as the bell-MEPPs. Unitary EPPs were recorded from neonatal and re-innervating preparations by reducing the evoked response with cobalt ions (4 mM). Distributions of unitary EPPs were similar to those of bell-MEPPs. It is concluded that there are two classes of spontaneous quanta. The skew-MEPP class dominates MEPP amplitude distributions during the early stages of re-innervation and early neonatal preparations. In all stages of development the unitary evoked EPPs have the same mean amplitude and time-to-peak as the bell-MEPPs. The data suggest that the skew class is not available for evoked release.