Paola Paggi

The maximum rate of neurofilament transport in axons: a view of molecular transport mechanisms continuously engaged

Neurofilaments (NFs) were radiolabeled in the optic systems of mice. The leading edge of the radiolabeled NF waveform was distinguished near the injection site (the eye) both by liquid scintillation spectroscopy and visually from fluorographs. The fastest NFs were found to be translocated at rates of between 72 and 144 mm/day. It appears that the continuous (maximal) operation of the slow axonal transport machinery can move polymers intra-axonally at rates one hundred times greater than those previously reported.

Effect of Latrodectus mactans tredecimguttatus venom on the crayfish stretch receptor neurone

Abstract Latrodectus venom is toxic to the crayfish Astacus astacus , its toxicity being in the same range as that for the guinea pig and housefly. Low doses of the venom affect the activity of the stretch receptor neuron of the crayfish modifying and blocking the impulse frequency. Picrotoxin does not prevent this effect.

Effect of Latrodectus mactans tredecimguttatus venom on sympathetic ganglion isolated in vitro.

Abstract The effect of Latrodectus venom (L.V.) on the function of the superior cervical ganglion of the rat, isolated in vitro , has been studied. This venom promptly depresses the postganglionic action potentials evoked by preganglionic stimulation, while it elicits discharges of asynchronous action potentials from the unstimulated ganglion. Previous curarization effectively interferes with the production of these discharges. When unstimulated ganglion (previously incubated with H 3 -choline for 2 hr of repetitive activity) was treated with L.V., a 50 per cent decrease in its labelled acetylcholine content (relative to the control ganglion) was observed

Gene expression pathways induced by axotomy and decentralization of rat superior cervical ganglion neurons

To identify genes potentially involved in remodelling synaptic connections, we induced the temporary detachment of pre‐ and post‐synaptic elements by axotomy or denervation of rat superior cervical ganglion neurons. cDNA microarray analysis followed by stringent selection criteria allowed the identification of a panel of genes whose expression was modulated by axotomy at various time points after injury. Among these genes, 11 were validated by real‐time reverse transcriptase‐polymerase chain reaction on independently prepared samples after superior cervical ganglion neuron axotomy (1, 3 and 6 days) and compared with the effect of decentralization (8 h, 1 and 3 days). These genes code for extracellular matrix/space [apolipoprotein D (apoD), decorin, collagen alpha1 type I, collagen alpha1 type III] and intermediate filament (vimentin) proteins, for modulators of neurite outgrowth (thrombin receptor, plasminogen activator inhibitor‐1, bone morphogenetic protein 4, annexin II and S‐100‐related protein, clone 42C) and for a nerve cell transcription factor (brain finger protein). Eight of these 11 genes showed significant and persistent modulations after both types of injury. Finally, protein levels of apoD were shown to increase in superior cervical ganglion after axotomy. Our results identify hitherto unrecorded genes responsive to axotomy and decentralization of superior cervical ganglion neurons, and probably involved in synapse formation, remodelling and elimination.

Dystrophin Stabilizes α3- But Not α7-Containing Nicotinic Acetylcholine Receptor Subtypes at the Postsynaptic Apparatus in the Mouse Superior Cervical Ganglion

The nicotinic acetylcholine receptor (nAChR) subtypes were characterized in the superior cervical ganglion (SCG) of wild-type and dystrophin-lacking mdx mice. The binding of Epibatidine and alphaBungarotoxin, ligands for alpha3- and alpha7-containing receptors, respectively, revealed, for each ligand, a single class of high-affinity binding sites, with similar affinity in both wild-type and mdx mice. The Epibatidine-labeled receptors were immunoprecipitated by antibodies against the alpha3, beta2, and beta4 subunits. Immunocytochemistry showed that the percentage of alpha3-, beta2-, and beta4- but not of alpha7-immunopositive postsynaptic specializations was significantly lower in mdx than in wild-type mouse SCG. These observations suggest that the mouse SCG contains nAChRs, stabilized by dystrophin, in which the alpha3 subunit is associated with the beta2 and/or beta4 subunits. Conversely, dystrophin is not involved in the stabilization of the alpha7-containing nAChRs, as the percentage of alpha7-immunopositive synapses is similar in both wild-type and mdx mouse SCG.

Dystrophin and its isoforms in a sympathetic ganglion of normal and dystrophic mdx mice : Immunolocalization by electron microscopy and biochemical characterization

In normal mouse superior cervical ganglion, dystrophin immunoreactivity is present in ganglionic neurons, satellite cells and Schwann cells. It is associated with several cytoplasmic organelles and specialized plasma membrane domains, including two types of structurally and functionally different intercellular junctions: synapses, where it is located at postsynaptic densities, and adherens junctions. Dystrophin immunostaining can be ascribed to the 427,000 mol. wt full-length dystrophin, as well as to the several dystrophin isoforms present in superior cervical ganglion, as revealed by western immunoblots. In mdx mouse superior cervical ganglion, which lacks the 427,000 mol. wt dystrophin, the unchanged pattern of dystrophin immunolabelling observed at several subcellular structures indicates the presence of dystrophin isoforms at these sites. Moreover, the absence of labelled adherens junctions indicates the presence of full-length dystrophin at this type of junction in the normal mouse superior cervical ganglion. The lower number of immunopositive postsynaptic densities in mdx mouse superior cervical ganglion than in normal mouse ganglion suggests the presence, in the latter, of postsynaptic densities with differently organized dystrophin cytoskeleton: some containing dystrophin isoforms alone or together with 427,000 mol. wt dystrophin, and others containing 427,000 mol. wt dystrophin alone.

Neuronal compartments and axonal transport of synapsin I

Studies on the transport kinetics and the posttranslational modification of synapsin I in mouse retinal ganglion cells were performed to obtain an insight into the possible factors involved in forming the structural and functional differences between the axon and its terminals. Synapsin I, a neuronal phosphoprotein associated with small synaptic vesicles and cytoskeletal elements at the presynaptic terminals, is thought to be involved in modulating neurotransmitter release. The state of phosphorylation of synapsin I in vitro regulates its interaction with both synaptic vesicles and cytoskeletal components, including microtubules and microfilaments. Here we present the first evidence that in the mouse retinal ganglion cells most synapsin I is transported down the axon, together with the cytomatrix proteins, at the same rate as the slow component b of axonal transport, and is phosphorylated at both the head and tail regions. In addition, our data suggest that, after synapsin I has reached the nerve endings, the relative proportions of variously phosphorylated synapsin I molecules change, and that these changes lead to a decrease in the overall content of phosphorus. These results are consistent with the hypothesis that, in vivo, the phosphorylation of synapsin I along the axon prevents the formation of a dense network that could impair organelle movement. On the other hand, the dephosphorylation of synapsin I at the nerve endings may regulate the clustering of small synaptic vesicles and modulate neurotransmitter release by controlling the availability of small synaptic vesicles for exocytosis.

Effects of acute and repeated daily exposure to hypergravity on spatial learning in mice

Studies in humans have revealed that exposure to altered gravity may lead to impairments in cognitive functions. The objective of this study was to test whether mice exposed to hypergravity using a centrifuge apparatus showed learning impairments in a spatial learning task. Mice rotating at 1G or at 2G acceleration gravity and non-rotating controls were tested for reactivity to a spatial change after either a single 1 h or five repeated 1 h daily rotations in the centrifuge. While no differences among groups were found in the performance after single exposure to altered gravity, 5 days of repeated exposures to 1G or 2G gravity conditions significantly affected mouse ability to discriminate a new spatial arrangement. Additionally, this effect was stronger in the animals repeatedly exposed to 2G rather than to 1G conditions.

Effects of chromatographic fractions of black widow spider venom on in vitro biological systems.

Abstract Extracts of Latrodectus mactans tredecimguttatus venom glands were fractionated by means of gel filtration and ion exchange chromatography. Three protein peaks toxic to mice were assayed in vitro on three different preparations known to be affected by whole venom. The fractions elicited: (a) increased release of acetylcholine from rat brain cortex slices, (b) disappearance of the histochemical reaction due to catecholamines in adrenergic fibres of rat iris, and (c) amplitude decrease of preganglionically evoked action potentials recorded postganglionically from rat superior cervical ganglion, and appearance of asynchronous postganglionic action potentials recorded from unstimulated ganglia. Fractions C and D were active at protein concentrations as low as 2 μg/ml, while fraction B was considerably less potent.

Effects of denervation and lack of calcium on the action of Latrodectus venom on rat sympathetic ganglion.

Abstract Black-widow spider or Latrodectus mactans venom (L.V.) is known to block transmission of impulses in the isolated cord of the cockroach (1, 2) as well as the activity of the crayfish stretch receptor (3). It appears that, at least in the crayfish, this block is due to a direct depolarizing action of the venom on the somadendritic membrane (Obara and Mauro, personal communication). In contrast, L.V. blocks neuromuscular transmission in the frog, where, by acting presynaptically, it causes a large increase in the frequency of miniature end-plate potentials (MEPP), eventually leading to their exhaustion (4). A similar effect of L.V. on the neuromuscular junction was observed in the cat by Okamoto et al. (5) with no significant change in the muscular response evoked by direct electrical stimulation or by intra-arterial injection of acetylcholine (ACh.) When the venom is applied to isolated sympathetic ganglion of the rat it causes action potentials in the postganglionic cells and blocks synaptic transmission (6). It appears that this blocking action of the venom is again due to release and “exhaustion” of transmitter, since a) a previous curarization of the ganglion prevents the ganglion cell discharges and b) the venom causes a 50 per cent decrease in the labelled ACh content of the resting unstimulated ganglion which has been previously incubated with H3-choline for 2 hr while undergoing repetitive stimulation (6). The present study is a further attempt to elucidate the mode of action of L.V. First we have sought further evidence for the presynaptic action by studying its effect on previously denervated ganglia. A second approach has considered the effect on ganglia incubated in a Ca-free medium. This condition is known to prevent the release of ACh normally caused by presynaptic depolarization due to a nerve impulse or to a high-K medium (7). In fact, Longenecker et al. (4) reported that Ca2+ ions are not necessary in obtaining the effect of L.V. at the frog neuromuscular junction (viz. increased frequeency of MEPP).