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PubMed1995

Properties of synaptic transmission at single hippocampal synaptic boutons.

Synaptic transmission between individual presynaptic terminals and postsynaptic dendrites is a fundamental element of communication among central nervous system neurons. Yet little is known about evoked neurotransmission at the level of single presynaptic boutons. Here we describe key functional characteristics of individual presynaptic boutons of hippocampal neurons in culture. Excitatory postsynaptic currents (e.p.s.cs) were evoked by localized application of elevated K+/Ca2+ solution to single functional boutons, visually identified by staining with the vital dye FM1-43 (refs 6, 7). Frequent repetitive stimulation produced a decline in the incidence of e.p.s.cs as the pool of releasable vesicles was exhausted; typically, recovery proceeded with a time constant of about 40 s (23 degrees C), and involved a vesicular pool capable of generating about 90 e.p.s.cs without recycling. At individual synapses, synaptic currents were broadly distributed in amplitude, but this distribution was remarkably similar at multiple synapses on a given postsynaptic neuron. The average size of synaptic currents and of responses to focal glutamate application varied fourfold across different cells, decreasing markedly with increasingly dense synaptic innervation. This raises the possibility of a very effective mechanism for coordinating synaptic strength at multiple sites throughout the dendritic tree.

Action Potentials↗
PubMed2003

Sustained Ca2+ entry elicits transient postsynaptic currents at a retinal ribbon synapse.

Night (scotopic) vision is mediated by a distinct retinal circuit in which the light responses of rod-driven neurons are faster than those of the rods themselves. To investigate the dynamics of synaptic transmission at the second synapse in the rod pathway, we made paired voltage-clamp recordings from rod bipolar cells (RBCs) and postsynaptic AII and A17 amacrine cells in rat retinal slices. Depolarization of RBCs from -60 mV elicited sustained Ca2+ currents and evoked AMPA receptor (AMPAR)-mediated EPSCs in synaptically coupled amacrine cells that exhibited large, rapidly rising initial peaks that decayed rapidly to smaller, steady-state levels. The transient component persisted in the absence of feedback inhibition to the RBC terminal and when postsynaptic AMPA receptor desensitization was blocked with cyclothiazide, indicating that it reflects a time-dependent decrease in the rate of exocytosis from the presynaptic terminal. The EPSC waveform was similar when RBCs were recorded in perforated-patch or whole-cell configurations, but asynchronous release from RBCs was enhanced when the intraterminal Ca2+ buffer capacity was reduced. When RBCs were depolarized from -100 mV, inactivating, low voltage-activated (T-type channel-mediated) Ca2+ currents were evident. Although Ca2+ influx through T-type channels boosted vesicle release, as reflected by larger EPSCs, it did not make the EPSCs faster, indicating that activation of T-type channels is not necessary to generate a transient phase of exocytosis. We conclude that the time course of vesicle release from RBCs is inherently transient and, together with the fast kinetics of postsynaptic AMPARs, speeds transmission at this synapse.

Amacrine Cells↗
PubMed1993

Facilitation and depression at different branches of the same motor axon: evidence for presynaptic differences in release.

This study provides evidence that a neuron can exhibit differences in activity-dependent transmitter release at two synaptic sites due to variations in the properties of its presynaptic terminals. Two muscles in the stomatogastric system of the lobster Homarus americanus are innervated by a single motor neuron but respond differently to that motor neuron's input, resulting in two different movements evoked by one motor neuron. During continued motor neuron stimulation, the gm8 muscle contracts slowly and maintains contraction, while the gm9 muscle contracts rapidly and then relaxes. These different muscle responses can be accounted for, in large part, by the properties of the respective neuromuscular synapses: the excitatory junctional potentials recorded in gm8 are initially small but summate and facilitate with repeated stimulation, while those in gm9 are initially large but depress with repeated stimulation. Presynaptic differences in neurotransmitter release contribute strongly to the divergent responses; reduction of the excitatory junction potential amplitude by partial postsynaptic receptor blockade or by desensitization does not change the amount of depression at gm9. However, reduction of neurotransmitter release with low-Ca2+, high-Mg2+ saline removes gm9 synaptic depression and reveals that both neuromuscular junctions exhibit frequency-dependent homosynaptic facilitation. Postsynaptic differences in muscle input resistance and muscle composition may enhance the effects of the divergent release properties, but are not responsible for the activity-dependent changes. Ultrastructural features of the nerve terminals on the two muscles are consistent with the differential output of the terminals; the synapses on gm9 are larger and have more presynaptic dense bars than their counterparts on gm8. These data suggest that the basis for the differences in transmitter release between the two muscles may be a higher density of release sites in the gm9 synapses that leads to a higher output of neurotransmitter, rapid depletion of transmitter stores, and synaptic depression.

Aging↗
PubMed1994

Reinnervation of Pacinian corpuscles by CNS axons after transplantation to the dorsal column: incidence and ultrastructure.

We have investigated the capacity of injured axons in the spinal dorsal columns of young adult rats to reinnervate grafted Pacinian corpuscles. A branch of the hindlimb interosseous nerve with a group of crural Pacinian corpuscles attached to it was autotransplanted to the surface of the spinal cord and the nerve stump was implanted into the dorsal column. Two to three months later 16 grafts were removed for examination by light and electron microscopy. By 3 months after transplantation almost all Schwann cell columns of the grafted nerve branch were occupied by regenerated myelinated and unmyelinated axons. Of 41 corpuscles examined by electron microscopy 24 were reinnervated by 1-3 myelinated fibres which gave rise to multiple terminals in the inner core. The remaining corpuscles appeared to be denervated. Only two of the reinnervated corpuscles contained regenerated endings which reiterated the distinct ultrastructure of normal presynaptic terminals of CNS axons, characterized by clusters of lucent vesicles and paramembranous densities. All other corpuscles were reinnervated by terminals which resembled peripheral mechanosensory endings as they contained mitochondria and very few vesicles. One such corpuscle was coinnervated by small terminals filled with large dense cored vesicles. We assume that the majority of grafted Pacinian corpuscles have been reinnervated by dorsal column axons and that the regenerated terminals with the ultrastructure of peripheral mechanosensory endings derive from central axons of primary sensory neurons, which are apparently capable of constructing mechanosensory-like terminals in response to signals from the Pacinian corpuscles. The vesicle-filled endings are probably formed by second order sensory neurons, corticospinal neurons and small peptidergic neurons unable to adjust their terminals to the new target.

Animals↗
PubMed1988

Programmed cell death: cytochemical and X-ray microanalytical characterization of calcium compartments in neuromuscular junctions during the normal breakdown of the intersegmental muscles in the giant silkmoth Antheraea polyphemus.

Calcium stores were cytochemically demonstrated using a combined oxalate-pyroantimonate method in the neuromuscular junctions of the degenerating intersegmental muscles in the giant silkmoth Antheraea polyphemus. The elemental composition of punctate precipitates of the reaction product was determined by electron probe X-ray microanalysis of unstained thin sections by energy-dispersive spectrometry and wavelength-dispersive spectrometry. The wavelength-dispersive spectra collected over terminal axons demonstrate a significant calcium signal and a trace of antimony. During the rapid lytic phase of spontaneous muscle degeneration, the calcium punctate deposits were detected in presynaptic terminals in the following sites: the synaptic vesicles and the mitochondria. Calcium precipitates were also found in the dense bodies and the mitochondria encountered in the glial convolutions. No calcium deposit was seen in the synaptic clefts and intercellular spaces of the subsynaptic reticulum of type I and type II. A comparison of calcium to antimony ratios between the terminal axons and the sarcoplasmic lysosomes revealed highly significant differences (P less than 0.001). Such a variability of the calcium to antimony ratio may be related to different conditions of precipitation or antimony diffusion in the different cell compartments. It was concluded that such synaptic terminals do not appear damaged in spite of the muscle degeneration and presumably continue to perform vital functions while the muscles are no longer contractile 20 h after adult ecdysis.

Animals↗
PubMed1995

Distribution of glutamate receptor subtypes in the vertebrate retina.

The distribution of glutamate receptor subunit/subtypes in the vertebrate retina was investigated by immunocytochemistry using anti-peptide antibodies against AMPA (GluR1-4), kainate (GluR6/7) and metabotropic (mGluR1 alpha) receptors. All receptor subtypes examined are present in the mammalian retina, but they are distributed differentially. GluR1 is present in the inner plexiform layer as well as amacrine and ganglion cell bodies. GluR2 is present mainly in the outer plexiform layer and bipolar cells. An anti-GluR2/3 antibody labels both plexiform layers and various cell bodies in the inner nuclear layer and the ganglion cell layer. GluR4 is present on Müller glial cells. In the goldfish retina, GluR2 immunoreactivity is prominent in the Mb type of ON-bipolar cells, including the dendrites and the large synaptic terminal. The putative dendritic localization is surprising, because no depolarizing conductance increase induced by glutamate is thought to be present in these cells. An AMPA receptor at a presynaptic terminal is also unusual, and probably provides feedback control of glutamate release. GluR6/7 is most widespread in the retina, being present in horizontal, bipolar, amacrine and ganglion cells. Ion channels composed of GluR6 are now known to be phosphorylated by protein kinase A, resulting in current potentiation. This property and our present observation together suggest that the glutamate receptors previously studied electrophysiologically by others in horizontal cells may contain GluR6. mGluR1 alpha is found mostly in the inner plexiform layer; its localization partially overlaps with that of the inositol trisphosphate receptor in the retina. Our results suggest that, in the retina, glutamate receptor subtypes may be expressed in selective cell types according to their specific functions.

Animals↗
PubMed2004

In vivo imaging of neuronal activity by targeted expression of a genetically encoded probe in the mouse.

Genetically encoded probes show great promise in permitting functional imaging of specified neuronal populations in the intact nervous system, yet their in vivo application has been limited. Here, we have targeted expression of synapto-pHluorin, a pH-sensitive protein that reports synaptic vesicle fusion, to olfactory sensory neurons in mouse. Synapto-pHluorin selectively labeled presynaptic terminals of sensory neurons in glomeruli of the olfactory bulb. Odorant stimulation evoked large-amplitude fluorescence increases that were localized to individual glomeruli in vivo, correlated with presynaptic calcium influx, graded with stimulus intensity, and stable over a period of days. Spatial patterns of odorant-activated glomeruli were distributed and did not change systematically with increasing carbon chain length, in contrast to the finely organized chemotopy that has been reported using other imaging methods. Targeted expression of synapto-pHluorin in mouse will permit the analysis of previously inaccessible neuronal populations and chronic imaging from genetically identified neurons in vivo.

Aldehydes↗
PubMed1988

Role of calcium-activated potassium channels in transmitter release at the squid giant synapse.

1. Several compounds known to block Ca2+-activated K+ channels were microinjected into squid 'giant' presynaptic terminals to test the hypothesis that these channels mediate Ca2+-dependent neurotransmitter release. 2. Injection of tetrapentylammonium, nonyl-triethylammonium and decamethonium all reversibly blocked transmission evoked by presynaptic action potentials. 3. All three of these compounds blocked presynaptic Ca2+ channels. The actions of tetrapentylammonium on presynaptic Ca2+ influx were examined in detail and found to be quantitatively sufficient to account for the ability of this compound to inhibit transmitter release. 4. Injection of Ba2+, another agent known to block Ca2+-activated K+ channels, also reversibly blocked evoked transmitter release. Ba2+ simultaneously enhanced basal (asynchronous) transmitter release and thus may be decreasing evoked release by depleting transmitter quanta available for release. 5. None of these results provide any support for the hypothesis that Ca2+-activated K+ channels mediate Ca2+-dependent release of transmitter at the squid synapse. However, our results have identified a new class of compounds that block Ca2+ channels from their cytoplasmic surface.

Animals↗
PubMed1966

The action of calcium on neuronal synapses in the squid.

1. The isolated stellate ganglion of the squid (L. pealii) was studied with intracellular and extracellular micro-electrodes. Three or four nerve fibres in the preganglionic nerve establish synaptic relations with the giant axon in the last stellar nerve. Accordingly, 1-3 small presynaptic spikes (< 1 mV) could be recorded from within the post-synaptic axon.2. A micro-electrode was inserted in the presynaptic fibre and used to polarize and record simultaneously. In the distal (giant) synapse, hyperpolarization of the ending produced an increase in the size of the presynaptic action potential and post-synaptic potential (PSP). Depolarization had the opposite effect. These effects of polarization took more than 10 sec to develop fully, and declined with a similar time course at the end of polarization. Analogous results were obtained with two other preganglionic fibres, which make contacts in the proximal synaptic region.3. The second of a pair of preganglionic impulses evoked a PSP larger than the first. This facilitation of PSP was sometimes accompanied by a small increase in the size of the second action potential in the presynaptic axon. At some shorter intervals, the second presynaptic action potential was reduced in amplitude, but the PSP was still increased. Hyperpolarization of the presynaptic terminal increased the size of both PSPs in a pair and abolished the facilitation. With stronger hyperpolarization the second PSP was even smaller than the first.4. Removing or reducing the Ca in the bathing fluid reversibly abolished the post-synaptic response. The small presynaptic spikes remained practically unaffected. In these conditions a nerve impulse still invaded the ending and normal action potentials could be recorded from the pre-synaptic terminal. This shows that electrical coupling between pre- and post-synaptic axons is insufficient to account for synaptic transmission.5. In low-Ca solution synaptic transmission could be restored locally by extracellular ionophoretic application of Ca to a small portion of the synapse. At sensitive spots a post-synaptic current (recorded with the Ca pipette) and PSP could be detected earlier than 1 sec after commencing the application of Ca.6. Ca was ineffective when injected intracellularly into the presynaptic fibre at a spot where extracellular ionophoresis of Ca restored the PSP.7. The results indicate that synaptic transmission in the squid stellate ganglion is not electrical but due to the release of an unidentified transmitter. Release of this transmitter by the presynaptic nerve impulse requires the presence of Ca in the external medium. During the impulse Ca would combine with a ;Ca-receptor' in the membrane and initiate the reactions which lead to transmitter release. It appears that the ;Ca-receptor' is only accessible from the outside of the membrane.

Animals↗
PubMed1998

A neuronal correlate of secondary hyperalgesia in the rat spinal dorsal horn is submodality selective and facilitated by supraspinal influence.

Tissue injury produces hyperalgesia not only in the injured area (primary hyperalgesia) but also outside of it (secondary hyperalgesia). In the present investigation, the submodality selectivity and the contribution of supraspinal influence to a neural correlate of the secondary hyperalgesia induced by neurogenic inflammation was studied in the presumed pain relay neurons of the rat spinal dorsal horn. Mechanically and thermally evoked responses to wide-dynamic range (WDR) neurons of the spinal dorsal horn were recorded under sodium pentobarbital anesthesia in rats. Neurogenic inflammation was induced by application of mustard oil outside of the receptive fields of WDR neurons. To study the contribution of supraspinal influence to mustard oil-induced changes in neuronal responses, the spinal cord was transected at a midthoracic level or lidocaine was microinjected into the rostroventromedial medulla (RVM). Furthermore, the antidromically evoked compound volley in the sural nerve was determined to reveal excitability changes in the central terminals of primary afferent A-fibers induced by mustard oil. The results indicate that mustard oil adjacent to the receptive fields of spinal WDR neurons significantly enhanced their responses to mechanical but not to noxious heat stimuli, without a significant influence on their spontaneous activity. Both high- and low-threshold mechanoreceptive input to WDR neurons was equally facilitated, whereas mechanoreceptive input to spinal dorsal horn neurons mediating innocuous messages (low-threshold mechanoreceptive neurons) was not changed. Mustard oil in a remote site (forepaw) did not produce any hyperexcitability to responses evoked by hindpaw stimulation. Spinal transection or lidocaine block of the RVM significantly attenuated the mustard oil-induced mechanical hyperexcitability in spinal dorsal horn neurons. Mustard oil had no significant effect on a compound volley in the sural nerve induced by intraspinal stimulation of sural nerve terminals at a submaximal intensity. The selective mechanical hyperexcitability in spinal WDR neurons, without a change in their spontaneous activity, can be explained by a heterosynaptic facilitatory action on presynaptic terminals mediating mechanical signals to these nociceptive spinal neurons. These findings indicate that brain stem-spinal pathways, involving the RVM, do not only suppress nociception but under some pathophysiological conditions concurrent facilitatory influence may predominate and lead to enhancement of mechanical hyperexcitability. The descending facilitatory feed-back loop to nociceptive spinal neurons may help to protect the wounded tissue and thus promote healing.

Afferent Pathways↗
PubMed1994

Characterization of two central AMPA-preferring receptors having distinct location, function and pharmacology.

The existence of a pharmacological heterogeneity among the glutamate receptors sensitive to (RS)-alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) has been investigated in the adult rat central nervous system (CNS). AMPA stimulated [3H]noradrenaline ([3H]NA) release from hippocampal synaptosomes (pD2 = 4.58) and the production of cGMP in cerebellar slices (pD2 = 7.75). The AMPA effects in the two systems were tested against several glutamate receptor antagonists including 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), 6,7-dinitro-quinoxaline-2,3-dione (DNQX), L-glutamate diethylester (GDEE), gamma-D-glutamyl-glycine (GDGG) and gamma-D-glutamyl-aminomethylsulphonate (GAMS). In both systems the AMPA effect was equally sensitive to CNQX or DNQX. However, while the AMPA-evoked increase of [3H]NA release from presynaptic terminals was not affected by GAMS, GDGG or GDEE, the postsynaptic cGMP response was prevented by GDGG and GDEE. It is concluded that rat hippocampus and cerebellum possess, respectively, presynaptic and postsynaptic AMPA-sensitive receptors involved in different functions and endowed with diverse pharmacological properties.

Animals↗
PubMed1981

Morphological identification of axo-axonic and dendro-dendritic synapses in the rat substantia gelatinosa.

Axo-axonic and dendro-dendritic synapses of the rat substantia gelatinosa Rolandi (SGR) have been studied in 14 adult rats by means of thin section and freeze-etch electron microscopy. Out of 6045 synaptic contacts we identified 54 between axon terminals, 10 between dendritic processes. Aside from vesicle shape the following parameters were used in a morphometric study; width of synaptic clefts, depth and length of postsynaptic densities. There are two main types of axo-axonic synapses, those with agranular vesicles (AA-type) and those with an increased population of large granular vesicles (AAgr-type). The former is prevailing and can be subdivided into two subgroups: AA1 and AA2. The pre- and postsynaptic terminals of AA1 contain spherical vesicles, the depth and length of postsynaptic densities as well as the width of the junctional clefts being significantly larger than that of AA2. Presynaptic terminals of AA2 synapses contain predominantly flattened vesicles, while spherical vesicles were found exclusively in the postsynaptic boutons. AAgr-type is rarely encountered; it is characterized by large granular vesicles (65-110 nm) which accumulate postsynaptically and occasionally in both pre- and postsynaptic boutons. Two types of dendro-dendritic synapses (DD1 and DD2) constitute another SGR feature. The difference between DD1 and DD2 is analogous to that between AA1 and AA2 except that pleomorphic synaptic vesicles appear in both groups. The width of subsynaptic membrane appositions turned out to be the most consistent criterion by which the two groups could be differentiated.

Animals↗
PubMed1999

Synaptic density in geniculocortical afferents remains constant after monocular deprivation in the cat.

Monocular eyelid closure in cats during a critical period in development produces both physiological plasticity, as indicated by a loss of responsiveness of primary visual cortical neurons to deprived eye stimulation, and morphological plasticity, as demonstrated by a decrease in the total length of individual geniculocortical arbors representing the deprived eye. Although the physiological plasticity appears maximal after 2 d of monocular deprivation (MD), the shrinkage of deprived-eye geniculocortical arbors is less than half-maximal at 4 d and is not maximal until 7 d of deprivation, at which time the deprived arbors are approximately half their previous size. To study this form of plasticity at the level of individual thalamocortical synapses rather than arbors, we developed a new double-label colocalization technique. First, geniculocortical afferent arbors serving either the deprived or nondeprived eye were labeled by injection of the anterograde tracer Phaseolus vulgaris leucoagglutinin into lamina A of the lateral geniculate nucleus. Then, using antibodies to synaptic vesicle proteins, we identified presynaptic terminals within the labeled arbors in layer IV of the primary visual cortex. Analysis of serial optical sections obtained using confocal microscopy allowed measurement of the numerical density of presynaptic sites and the relative amounts of synaptic vesicle protein in geniculocortical afferents after both 2 and 7 d of MD. We found that the density of synapses in geniculocortical axons was similar for deprived and nondeprived afferents, suggesting that this feature of the afferents is conserved even during periods in which synapse number is reduced by half in deprived-eye arbors. These results are not consistent with the hypothesis that a rapid loss of deprived-eye geniculocortical presynaptic sites is responsible for the prompt physiological effects of MD.

Aging↗
PubMed1993

[An electron microscopic observation on spike foci of human epileptic brains].

Twenty cases of ECOG spike foci in epilepsy originally recorded by flaky silver electrodes on the surface of cortices and then to be removed surgically were studied under transmission electron microscope. The ultrastructural changes of the spike foci seemed to be identical in all cases although there were differences in etiologies, locations and seizure types. The presynaptic terminals of axospinal asymmetric synapses were obviously swollen anyhow, the postsynaptic spines were relatively unaffected. The synaptic vesicles were decreasing in number and to be accumulated predominantly at the presynaptic membrane which indicated that these excitatory changes of synapses may play an important role in the mechanism of epileptic abnormal electric activity and the development of epilepsy. Additionally, the ultrastructural changes and the cytopathologic significance of neurons, astrocytes and capillaries are also discussed.

Adolescent↗
PubMed1996

Selective, activity-dependent uptake of histamine into an arthropod photoreceptor.

The synapses made by many arthropod photoreceptors are disinhibitory and use histamine as their transmitter. Because decreases and not increases in the cleft concentration of transmitter constitute the important event at these synapses, a transporter to clear the cleft of histamine would seem particularly crucial to signal transfer. We report here that 3H-histamine is taken up selectively into barnacle photoreceptors by a Na+-dependent mechanism, presumably a transporter. Using light microscopic autoradiography, we observe heavy label over axons and presynaptic terminals of these neurons when they are stimulated during uptake. The radioactivity taken up was identified as 3H-histamine by thin layer chromatography; no metabolites were detected, even after 5 hr. Radiolabeled 5-hydroxytryptamine and GABA are not taken up by the photoreceptor. 3H-histamine uptake into photoreceptors is decreased markedly by an excess of unlabeled histamine and by chlorpromazine and phenoxybenzamine. Unexpectedly for uptake dependent on the NA+ gradient, photoreceptor terminals label more intensely in the light (when depolarized) than in the dark (when hyperpolarized). Glia label more strongly than photoreceptors in dark-incubated preparations. The presence of presynaptic uptake strengthens the evidence that histamine is the neurotransmitter of arthropod photoreceptors and provides a mechanism by which this synapse could recycle transmitter, control its steady-state cleft concentration, and clear it from the cleft in response to decreases in its release from the photoreceptors.

Animals↗
PubMed1997

Responses of cortical noradrenergic and somatostinergic fibres and terminals to adjacent strokes and subsequent treatment with NGF and/or the ganglioside GM1.

The occurrence of sprouting by fibre systems in the neocortex following lesion is still a controversial issue. In previous studies, we showed a nerve growth factor (NGF)-induced sprouting and hypertrophy of presynaptic terminals in the cholinergic fibres of the rat neocortex following stroke-type lesions, effects that were potentiated by the monosialoganglioside GM1. The present study investigated whether exogenous NGF and/or GM1 treatment could also affect the noradrenergic and somatostinergic systems in the neocortex. Immediately following unilateral vascular decortication, adult rats received, via minipump, a 7-day infusion of vehicle, NGF (12 microg/day) and/or GM1 (1.5 mg/day) into the cerebroventricular space. Thirty days postlesion, the animals were perfused with histological fixatives, the brains were removed, and relevant sections were processed for dopamine beta-hydroxylase and somatostatin immunocytochemistry at the light and electron microscopic levels. A Quantimet 920 image analysis system was used for the quantification of fibre length and size of presynaptic boutons. The lesion caused a reduction in the dopamine beta-hydroxylase-immunoreactive fibre length, which was not attenuated by either NGF or GM1 treatment or both. The somatostatin-immunoreactive network, in contrast, was unaffected by the lesion, and there was no sprouting of somatostatin fibres following trophic factor therapy. We also found no significant differences in the size and number of synapses of both the dopamine beta-hydroxylase-immunoreactive and somatostatin-immunoreactive boutons following lesion and drug treatments. These results indicate that NGF and/or GM1 therapies do not cause regrowth in the noradrenergic and somatostatinergic cortical fibre networks or their presynaptic elements following a cortical devascularizing lesion.

Analysis of Variance↗
PubMed1977

Quantitative autoradiographic light- and electron microscopic studies on the retinohypothalamic connections in the rat.

Light microscopic autoradiography performed subsequent to intraocular injection of 3H-leucine revealed silver grains (SG) above axons of the optic tract which could be followed into the ventral and caudal portion of the suprachiasmatic nuclei (SCN) and above the contralateral anterior hypothalamic nucleus (AHN). By high resolution photometric measurement and computer processing the labelled areas were analysed, thus yielding statistical data of the relative grain distribution. The highest SG density was found in the ventrolateral part of both SCN (SCvl), confirming earlier reports concerning retinohypothalamic connections. That area exhibiting a cytoarchitecture different from the remaining nucleus was traversed, however, by numerous labelled axons. In the caudal part of both SCN a specific projection field of retinal fibres could be located. Here, almost no traversing fibres contribute to the rather circumscribed marked area. In the ventral part of the contralateral AHN, diffuse labelling well above background levels could be observed. Distinction between bypassing and terminating fibres within the SCvl could not be made using light microscopy. Analysis of SG distribution of the SCvl with electron microscopic autoradiography revealed a specific localization of SG within presynaptic terminals containing clear vesicles and pale mitochondria.

Animals↗
PubMed1972

Protein synthesis in synaptosomal fractions. Ultrastructural radioautographic study.

A quantitative ultrastructural radioautographic study of in vitro protein synthesis has been carried out in rat synaptosomal fractions incubated with tritiated leucine or a tritiated amino acid mixture. Analysis of grain density distribution demonstrated that presynaptic endings are labeled. 30-50% of the developed grains, representing tritiated amino acids incorporated into proteins, were related to presynaptic endings which accounted for 75-77% of the total processes. 34-45% of the grains were related to processes containing ribosomes which accounted for only 4-7% of the total processes. The relative specific activity of these ribosome-containing processes, some of which could be identified as postsynaptic elements, was up to ten times higher than that of the presynaptic ending. These findings indicate that protein synthesis takes place in vitro in presynaptic terminals although to a significantly lesser degree than that occurring in ribosome-containing processes, which, with other nonpresynaptic processes, are at the present time unavoidable contaminants of synaptosomal fractions. Presynaptic endings that in radioautographs contained no mitochondria were labeled. Also, presynaptic endings were labeled after incubation in the presence of chloramphenical which inhibited 20% of the protein synthesis of the synaptosomal fraction. It is concluded that besides mitochondrial protein synthesis, another protein synthesizing system operates in presynaptic endings in vitro.

Amino Acids↗