Guy Massicotte

Mechanism of altered synaptic strength due to experience: relation to long-term potentiation

An increase in medial perforant synaptic strength can be observed for hippocampal slices from rats exposed to environmental enrichment. The expression of enhanced synaptic strength exhibits properties similar to long-term potentiation (LTP), a physiological model of memory storage. Similarities include an increase in strength of the synaptic response in the absence of an altered paired-pulse ratio and an increase in the binding of the glutamate agonist alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate. Furthermore, environmental enrichment interacts with the mechanisms responsible for the induction of LTP by inhibiting further increases in synaptic strength following LTP-inducing stimulation. The results provide evidence for experience-mediated influences on postsynaptic mechanisms regulating medial perforant path synaptic strength.

Impaired modulation of AMPA receptors by calcium-dependent processes in streptozotocin-induced diabetic rats.

The mechanisms by which diabetes impairs cognitive function are not well-established. In the present study, we determined the electrophysiological and biochemical nature of disturbances in the mechanism of long-term potentiation (LTP) in diabetic rats. As previously reported, the administration of streptozotocin (STZ) was found to reduce the magnitude of LTP in the CA1 region of the hippocampus, while the same treatment did not interact with the capacity of the hippocampus to generate long-term depression induced by low-frequency stimulation. In addition, STZ treatment did not modify the component of excitatory postsynaptic potentials mediated by activation of the N-methyl-D-aspartate (NMDA) subtype of glutamate receptors, suggesting that NMDA receptor function remained intact in STZ-treated slices. At the biochemical level, the capacity of calcium to increase [3H](RS)-alpha-amino-3-hydroxy-5-methylisoxazole propionic acid (3H-AMPA) binding to glutamate/AMPA receptors in rat brain tissue sections was markedly affected in most regions of the hippocampus of STZ-treated rats. Moreover, changes in 3H-AMPA binding properties elicited by both exogenous phospholipase A2 and melittin, a potent activator of endogenous phospholipases, were also altered in synaptoneurosomes from diabetic rats. Taken together, the present data suggest that the loss of LTP maintenance in STZ-treated rats is more likely the result of disruption of calcium-dependent processes that are suspected to modulate postsynaptic AMPA receptors during synaptic potentiation. Understanding the biochemical factors participating in the impairment of AMPA receptor modulation might provide important clues revealing the very basis of memory deficits in diabetes.

Effect of bromophenacyl bromide, a phospholipase A2 inhibitor, on the induction and maintenance of LTP in hippocampal slices

The effect of bromophenacyl bromide (BPB), a phospholipase A2 (PLA2) inhibitor, on both the induction and the maintenance of long-term potentiation (LTP) was investigated in field CA1 of the hippocampal slice preparation. One hour of BPB application (50 microM) caused a large reduction in the magnitude of LTP induced by a theta burst stimulation (TBS) paradigm. BPB had no significant effect on either the degree of paired-pulse facilitation or the amount of pre-established LTP. Furthermore, the facilitation of postsynaptic responses occurring during TBS and in the first minute following TBS was not reduced by the PLA2 inhibitor. These results indicate that the inhibition of LTP produced by BPB is not due to an effect of the drug on a physiological event that triggers LTP. The data also suggest that PLA2 activation plays a critical role in the expression of LTP, but is not required for the maintenance of the potentiation.

Modulation of dl-α-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA)/quisqualate receptors by phospholipase A2 treatment

The expression of long-term potentiation (LTP) in area CA1 of hippocampus has been proposed to result from an increased sensitivity of the AMPA/quisqualate receptors. We have investigated the binding properties of excitatory amino acid receptors in phospholipase A2 (PLA2)-treated rat brain membranes. PLA2 from bee venom produced a significant increase in the binding of [3H]-AMPA ([3H]-amino-3-hydroxy-5-methylisoxazole-4- propionate), a ligand for the AMPA/quisqualate receptor. Analysis of the saturation kinetics revealed that PLA2 treatment increased the affinity of the AMPA/quisqualate receptor without changing the maximum number of sites. In contrast, PLA2 treatment did not detectably modify the binding of [3H]-kainate to the kainate receptor and of [3H]-glutamate and [3H]-glycine to the NMDA (N-methyl-D-aspartate) receptor complex. These finding suggest that phospholipase A2 may regulate the AMPA/quisqualate receptor and could play an important role in the development of LTP.

CRMP3 is required for hippocampal CA1 dendritic organization and plasticity

In vitro studies have pointed to the collapsin response mediator proteins (CRMPs) as key regulators of neurite outgrowth and axonal differentiation. CRMP3 is expressed mostly in the nervous system during development but remains at high levels in the hippocampus of adults. To explore CRMP3 function in vivo, we generated mice with targeted disruption of the CRMP3 gene. Immunohistochemistry and Golgi staining of CA1 showed abnormal dendrite and spine morphogenesis in the hippocampus of CRMP3‐deficient mice. Apical dendrites displayed an increase in undulation and a reduction in length and branching points. Basal dendrites also exhibited a reduction in length with an alteration in soma stem distribution and an increased number of thick dendrites localized in stratum oriens (SO). Long‐term potentiation (LTP) was impaired in this area. These data indicate an important role for CRMP3 in dendrite arborization, guide‐posts navigation, and neuronal plasticity.—Quach, T. T., Massicotte, G., Belin, M. F., Honnorat, J., Glasper, E. R., Devries, A. C., Jakeman, L. B., Baudry, M., Duchemin, A. M., Kolattukudy, P. E. CRMP3 is required for hippocampal CA1 dendritic organization and plasticity. FASEB J. 22, 401–409 (2008)

Blunted Responses to Vasoconstrictors in Mesenteric Vasculature but not in Portal Vein of Spontaneously Hypertensive Rats Treated with Relaxin

Abstract Relaxin (RLX), an ovarian polypeptide hormone that is particularly associated with gestation in viviparous species, has recently been shown to decrease blood pressure in virgin spontaneously hypertensive rats (SHR) upon chronic infusion. In this investigation, vascular reactivity to angiotensin II, arginine-vasopressin, and norepinephrine was studied in the perfused mesenteric artery and isolated portal vein of control and RLX-treated virgin spontaneously hypertensive rats. The latter received an intravenous infusion of 75 ng/hr purified rat RLX for 2 days, whereas the controls were given an equal infusion of saline. All of the animals were then killed and their tissues processed for in vitro study. In the perfused mesenteric artery, the concentration-response curves for arginine-vasopressin and norepinephrine were shifted to the right by a factor of about 2 (P < 0.05 and P < 0.005, respectively) after RLX treatment. In the isolated portal vein, the response to angiotensin II was not affected; the effect of norepinephrine was slightly displaced to the right (increase in EC50) and the maximum response remained unchanged. These results demonstrate that RLX treatment for 42 hr blunted the vascular response to vasoconstrictor agents in the mesenteric vasculature and are consistent with similar observations reported previously in the same tissue of 20-day-old pregnant rats. It is concluded that RLX may be involved in the blunted response to vasoconstrictor agents during gestation in the rat.

AMPA receptor properties in adult rat hippocampus following environmental enrichment

In adult rats, environmental enrichment has been shown to selectively increase -AMPA binding in the hippocampus but the molecular mechanisms underlying this effect remain unknown. We used in situ hybridization with antisense oligonucleotides to determine possible changes in the hippocampal expression of messenger RNAs for different subunits of AMPA receptors in adult rats following exposure to an enriched environment. Quantitative analysis revealed that mRNA levels for three subtypes of AMPA glutamate receptors (GluR1-3; Flip and Flop variants) were not modified in any hippocampal region after environmental enrichment. In addition, no differences were detected in the levels of GluR1 and GluR2/3 proteins in Western blots of hippocampal membranes from enriched rats. Nevertheless, quantitative ligand binding autoradiography indicated that environmental enrichment evoked a significant and uniform decrease in the capacity of calcium or phosphatidylserine (PS) to up-regulate -AMPA binding in various hippocampal regions but not in the cerebral cortex. These findings support previous observations suggesting that post-translational changes in AMPA receptor properties, as a result of the activation of calcium-dependent processes, may represent an important mechanism underlying long-term modifications of synaptic efficacy in the rat hippocampus.

Bidirectional modulation of AMPA receptor properties by exogenous phospholipase A2 in the hippocampus

The synaptic modifications underlying long‐term potentiation (LTP) and long‐term depression (LTD) of synaptic transmission in various brain structures may result from changes in the properties of the α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazole propionate (AMPA) subtype of glutamate receptors. In the present study, we report that treatment of rat synaptoneurosomes with increasing concentrations of phospholipase A2 (PLA2) produces a biphasic effect on AMPA receptor binding, with low concentrations causing a decrease and high concentrations an increase in agonist binding. Analysis of the saturation kinetics of 3H‐AMPA binding revealed that the biphasic effect of PLA2 was due to modifications in receptor affinity and not to changes in the maximum number of binding sites for AMPA receptors. The 12‐lipoxygenase inhibitors preferentially reduced PLA2‐induced decrease in AMPA binding and treatment of hippocampal synaptoneurosomes with arachidonic acid (AA) or 12‐HPETE, the first metabolite generated from the hydrolysis of AA by 12‐lipoxygenases, decreased 3H‐AMPA binding. Moreover, electrophysiological experiments indicated that the 12‐lipoxygenase inhibitor baicalein totally blocked LTD formation in area CA1 of hippocampal slices. The decrease in 3H‐AMPA binding elicited by low concentrations of PLA2, as well as the level of LTD, were partially reduced by AA‐861, a 5‐lipoxygenase inhibitor, while the cyclooxygenase inhibitor indomethacin did not prevent LTD formation or the effects of PLA2 on 3H‐AMPA binding. Our results provide evidence for a possible involvement of lipoxygenase metabolites in the regulation of AMPA receptor during synaptic depression. In addition, they strongly support the idea that the same biochemical pathway, i.e., NMDA receptor activation and endogenous PLA2 stimulation, may represent a common mechanism resulting in AMPA receptor alterations for both LTP and LTD formation. Hippocampus 1998;8:299–309.

Dopamine D1 receptor activation induces tau phosphorylation via cdk5 and GSK3 signaling pathways.

Increasing evidence is demonstrating that drugs affecting dopamine levels in the brain induce cytoskeletal modifications. These evolving changes may impact neuronal synaptic plasticity as cytoskeletal constituents are involved in the maintenance of dendritic processes, and any alterations in their stability could influence major cellular compartments of neurons, such as dendrites, spines and synapses. Here, we describe a molecular chain of events that links dopamine D1 receptor activation to hyperphosphorylation of the microtubule-associated protein tau, which is normally involved in microtubules stabilization. We show, in SK-N-MC cells and rat striatal sections, that phosphorylation of tau at serines 199-202 and 214 appears to be mediated through activation of calcium-dependent intracellular mechanism, subsequent to D1 receptor-induced cAMP-dependent protein kinase A (PKA). We demonstrate, using pharmacological tools, that PKA activation causes increase of calcium levels, leading to cyclin-dependent kinase 5 activation by calpain proteolysis of p35 to p25 and glycogen synthase kinase 3beta activation by its phosphorylation at tyrosine 216. The D2 receptor agonism or lowering cAMP levels has no effect in our experimental settings. Moreover, we do not observe any association between phosphorylated tau and cellular damage. These data unravel novel mechanisms of tau hyperphosphorylation during G-protein-coupled receptor activation and are the first to show that stimulation of D1 receptors could have a profound influence on the neuronal cytoskeletal constituent tau.

Preserved LTP and water maze learning in hyperglycaemic-hyperinsulinemic ZDF rats

Previous investigations have demonstrated that cognitive deficits as well as hippocampal dysfunctions are generated in animals presenting manifestations of Type 1 diabetes (T1D) mellitus. The present study examined whether such deficits can also be reproduced in the Zucker Diabetic Fatty (ZDF) rats after they developed symptoms of Type 2 diabetes (T2D). Learning and memory assessments were performed using the Morris water maze 5 weeks after the animals presented symptoms of Type 1 diabetes for Experiment 1 (Exp 1) and after 8 weeks for Experiment 2 (Exp 2). Testing in the water maze revealed that ZDF rats learned the task normally, although control rats were found to swim significantly faster after 5 or 8 weeks of untreated diabetes. From an electrophysiological perspective, we observed that the integrity of synaptic function was also preserved in ZDF rats as no alterations in long-term potentiation (LTP) were observed in the area CA1 of hippocampal slices. It is concluded that hyperglycaemia is not the only factor influencing water maze learning and LTP in this animal model of Type 2 diabetes (T2D). The experiments suggest that the resistance of ZDF rats to cognitive and electrophysiological dysfunctions might be related to the protective action of hyperinsulinemia. Indeed, measurements of the plasma insulin level at the end of testing were significantly superior in ZDF rats in comparison to control rats.