Most recent publications
Cyclic guanosine-3′,5′-monophosphate (cyclic-GMP or cGMP) is a classical second messenger involved in the control of different physiological functions. cGMP is synthesised by the family of guanylyl cyclases that includes soluble and particulate enzymes, present in several isoforms and with different activation. cGMP effects are mainly mediated by PKG and CNG channels, whereas they are terminated by either specific or dual-substrate phosphodiesterases. In the central nervous system (CNS), cGMP has attracted the attention of neuroscientists especially for its key role in long-term potentiation, the synaptic plasticity phenomenon underlying memory formation and consolidation, thus setting off a “gold rush” for novel therapeutic approaches for the treatment of cognitive deficits. In this review, we have summarized the current knowledge on the neurochemistry of the cGMP system and the pre-clinical and clinical evidence on the use of cGMP enhancers in Alzheimer’s disease (AD) therapy. Although preclinical data demonstrates the beneficial effects of cGMP on cognitive deficits in AD animal models, the results of the clinical studies are rather inconclusive and, therefore, more trials with a dose-finding design on selected AD patient’s cohorts, possibly investigating also combination therapies, are still needed to evaluate the clinical potential of cGMP enhancers.
Di Giovanni G., Grandi L.C., Fedele E., Orban G., Salvadè A., Song W., Cuboni E., Stefani A., Kaelin-Lang A., Galati S. (2020) Acute and Chronic Dopaminergic Depletion Differently Affect Motor Thalamic Function. Int. J. Mol. Sci. 21(8). pii: E2734.
Although the motor thalamus (MTh) is known to play a key role in the codification of motor information through the basal ganglia (BG)-cortical loop, little is known with regard to its functional alterations in pathological states, such as Parkinson’s disease. To this purpose, in this study, we examined the effects of acute and chronic dopamine depletion on the neuronal firing of MTh neurons and on the cortical/MTh interplay by electrophysiology techniques. In addition, we have monitored MTh extracellular concentrations of glutamate and GABA by intracerebral microdialysis. We induced an acute dopamine depletion by infusing tetrodotoxin (TTX) into the medial forebrain bundle (MFB) in anesthetized rats. Chronic dopamine depletion was obtained by 6-hydroxydopamine (6-OHDA)-induced MFB degeneration. Acute dopamine depletion resulted in a significant inhibition of MTh neuronal activity without changes in burst content. On the contrary, chronic dopamine depletion did not change the firing rate but increased the burst firing, and affected the MTh-cortical coupling. As for neurotransmitter release in the MTh, acute dopamine depletion evoked a significant increase of extracellular GABA concentration and no change of glutamate levels, whereas we observed a significant reduction of extracellular GABA and an increase of glutamate when dopamine was chronically depleted. Our results show that the activity of MTh is indeed altered by DA depletion and support the hypothesis that, under chronic dopamine deprivation, a rebalancing of MTh function occurs, which is able to counterbalance the marked changes brought about by acute dopamine depletion.
Bonifacino T.*, Rebosio C.*, Provenzano F., Torazza C., Balbi M., Milanese M., Raiteri L., Usai C., Fedele E., Bonanno G. (2019) Enhanced function and overexpression of metabotropic glutamate receptors 1 and 5 in the spinal cord of the SOD1G93A mouse model of amyotrophic lateral sclerosis disease progression. Int. J. Mol. Sci. 20(18). pii: E4552. *equally contributed.
Glutamatergic excitotoxicity is one of the main causes of Amyotrophic Lateral Sclerosis (ALS) which leads to the death of primary and secondary motor neurons. In one of our previous preclinical studies, using SOD1G93A mice as a model of the human pathology, we showed that the metabotropic glutamatergic receptors of group I (mGluR1/5) were able to induce an abnormal release of glutamate in the spinal cord at a late stage of the disease (120 days). In order to understand whether this event could be the cause/concause of the disease or its effect, we conducted a series of experiments at the pre-symptomatic phase of the disease (30 and 60 days) or at the beginning of the clinical symptomatology (90 days). We observed that, in the spinal cord, the release of glutamate induced by 3,5-DHPG, a mixed mGluR1/5 agonist, was increased at 90 days in SOD1G93A mice, while at 30 and 60 days there were no differences with respect to control animals. The pharmacological characterization of this effects with selective antagonists revealed that both mGluR1 and mGluR5 were involved. Moreover, the expression of mGluR1 and mGluR5 on glutamatergic nerve endings was increased in the spinal cord of SOD1G93A mice at 90 days. Therefore, we can conclude that, in the early symptomatic phase, the function and expression of group I metabotropic glutamate receptors are increased in the spinal cord of SOD1G93A mice, causing a further enhancement of glutamate release that may participate in the neurodegenerative processes involved in ALS.
van Goethem N.P*., Paes D*., Puzzo D*., Fedele E*., Rebosio C., Gulisano W., Palmeri A., Wennogle L.P., Peng Y., Bertrand D., Prickaerts J. (2019) Antagonizing α7 nicotinic receptors with methyllycaconitine (MLA) potentiates receptor activity and memory acquisition. *equally contributed. Cell Signal. 62:109338, 1-8
It is well known that cholinergic neurotransmission is involved in cognitive processes. In particular, stimulation of α7-nicotinic receptors (α7-nAChRs) by selective agonists or positive allosteric modulators (PAM) has been proposed as a therapeutic approach for different neurological and psychiatric disorders. Nevertheless, there are no nicotinic ligands approved for clinical use. In this article, we report that low (picomolar) concentrations of the selective α7-nAChR antagonist methyllycaconitine (MLA) are able to potentiate electrophysiological responses of human α7-nAChRs and to convert long-term potentiation from the early form (E-LTP) to the late one (L-LTP) in the hippocampus. In addition, intraperitoneal administration of low doses of MLA were able to increase hippocampal glutamate extracellular levels in microdialysis experiments and to improve memory acquisition processes in the object recognition task (ORT). These results are at variance with the well known amnestic effects of nAChR antagonists that, however, manifest at higher doses. We hypothesise that low concentrations of MLA increase α7-nAChR activity by decreasing its desensitization state or by optimizing ACh binding to the receptor, thus maximally stabilizing channel opening.
These results indicate that, in addition to agonists and PAMs, α7-nAChR antagonists, at appropriate doses, may also be beneficial for the treatment of disorders characterized by cognitive deficits, such as Alzhemeir’s disease.
Rosenbrock H., Giovannini R., Schänzle G., Koros E., Runge F., Fuchs H., Marti A., Reymann K.G., Schröder U.H., Fedele E., Dorner-Ciossek C. (2019) The Novel Phosphodiesterase 9A Inhibitor BI 409306 Increases Cyclic Guanosine Monophosphate Levels in the Brain, Promotes Synaptic Plasticity, and Enhances Memory Function in Rodents. J. Pharmacol. Exp. Ther. 371(3):633-641
In this article, the novel and potent PDE9A inhibitor BI409306 has been pharmacologically characterized in terms of its potency (IC50 of 65 and 168 nM versus human and rat PDE9A, respectively) and selectivity (versus 9 different PDE isoforms) and its ability to increase cGMP, in vitro and in vivo, and to potentiate hippocampal LTP, the electrophysiological substrate of learning and memory formation. Moreover, the drugs has been shown to reverse deficits of working memory in the T-maze alternation test in mice treated with the NMDA receptor antagonist MK-801 and to enhance episodic memory in normal mice in the object recognition test with a 24-h delay schedule. Preliminary results on Alzheimer’s disease patients showed that BI 409306 was well tolerated but it did not show significant effects on cognitive deficits at the tested doses.
Olivero G, Vergassola M, Cisani F, Usai C, Pittaluga A. (2019) Immuno-Pharmacological Characterization of Presynaptic GluN3A-Containing NMDA Autoreceptors: Relevance to Anti-NMDA Receptor Autoimmune Diseases. Mol Neurobiol. 56(9): 6142-6155.
The study represents an extension of previous investigations aimed at deciphering the subunit composition of presynaptic NMDA autoreceptors in the hippocampus. Our previous results showed that GluN1, GluN2A, GluN2B and GluN3A subunits exist in the presynaptic component of hippocampal synaptosomal plasmamembranes, where they associate to form NMDA receptors (NMDAR, Musante et al., 2011, J Neurochem 117:516-527). The participation of the GluN1, GluN2A and GluN2B subunits was supported by functional results with selective GluN agonists and antagonists. The lack of selective ligands for the GluN3A subunit, on the contrary, did not allow to confirm the presence of this subunits, although few functional observations seemed predictive of its involvement in the NMDA receptor assembly. Owing to investigate this aspect, we adopted the “immune-pharmacological” approach, which consists on the use of antibodies raised against the amino-terminal sequence of a selected receptor subunit as selective tools to highlight the participation of the protein to the receptor assembly. By using this approach, we proved the participation of GluN3A subunit to the expression of presynaptic release-regulating NMDARs in hippocampal glutamatergic terminals. Our study also demonstrated the involvement of GluN1 subunit lacking the N1 cassette in the NMDAR assembly. We speculate that the glycinergic subunit permits the insertion of the GluN3A protein in synaptosomal plasmamembranes, favouring its association to a GluN1 / GluN2A / GluN2B heterodimeric complex. Finally, we demonstrated that the anti-GluN antibodies hamper the releasing activity elicited by NMDARs by triggering the internalization of NMDA receptor complex. Taking into account the role of anti-GluN autoantibodies in psychosis, our result addnew insights on the cellular events leading to disrupted synaptic plasticity in patients suffering of anti-NMDA-dependent psychiatric diseases.
Bonfiglio T, Olivero G, Vergassola M, Di Cesare Mannelli L, Pacini A, Iannuzzi F, Summa M, Bertorelli R, Feligioni M, Ghelardini C, Pittaluga A. (2019) Environmental training is beneficial to clinical symptoms and cortical presynaptic defects in mice suffering from experimental autoimmune encephalomyelitis. Neuropharmacology. 145(Pt A):75-86.
Environmental enrichment (EE) refers to the addition of objects to the animal’s cage to increase the levels of novelty and complexity. It increases cognitive activity and spontaneous physical exercise, potentiating synaptic plasticity to cope brain damage. EE is neuroprotective during aging and in a variety of neurodegenerative disorders and autoimmune demyelinating disease. In particular, early environmental training reduces the gravity of clinical score in mice suffering from Experimental Autoimmune Encephalomyelitis [EAE, an animal model used to study the pathophysiology of Multiple Sclerosis]. EE also increases the mobilization of adult neuronal progenitor cells into demyelinating lesions in these mice, also favouring the maturation of the stem cells from the subventricular zone to oligodendrocytes (Magalon et al., 2007, Eur. J. Neurosci. 25, 761). The study aimed at quantify the impact of “prophylactic” EE on clinical symptoms and presynaptic defects in EAE mice at the acute stage of disease (21 ± 1 days post immunization, d.p.i.). In EAE mice raised in an enriched environment (EE-EAE mice), the clinical score was reduced when compared to EAE mice raised in standard environment (SE-EAE). Concomitantly, gain of weight and increased spontaneous motor activity and curiosity were observed, suggesting increased well-being in mice. Impaired glutamate exocytosis and cyclic adenosine monophosphate (cAMP) production in cortical terminals recovered in EE-EAE mice when compared to SE-EAE mice. Finally, inflammatory infiltrates were reduced in the cortex but not in the spinal cord of EE-EAE mice while demyelination in the spinal cord was significantly reduced. We conclude that “prophylactic” EE is beneficial to synaptic derangements and preserves glutamate transmission in the cortex of EAE mice.
This review summarizes the main scientific evidence showing that the two cyclic nucleotides cAMP and cGMP, classic intracellular second messengers but with important extracellular functions, play a fundamental role in the physiological processes that underlie early and late long-term potentiation and, therefore, in the molecular and cellular mechanisms necessary for the formation of short- and long-term memory. But recent results, also obtained in the authors’ research laboratories, have shown that physiological levels of the beta amyloid peptide (Aβ), better known as one of the responsible for Alzheimer’s disease and the associated cognitive deficits, is indeed necessary for memory formation and consolidation, being the molecular effector of cAMP and cGMP promnesic activity. Based on these new data, the authors propose a new hypothesis that poses a loss of function of the Aβ peptide or in its reduced availability, and not its accumulation, as the possible cause of Alzheimer’s disease.
Rebosio C.*, Balbi M.*, Passalacqua M., Ricciarelli R., Fedele E. Presynaptic GLP-1 receptors enhance the depolarization-evoked release of glutamate and GABA in the mouse cortex and hippocampus. Biofactors. 2018, 44(2):148-157. *equally contributed.
The type 1 glucagon-like peptide (GLP-1) is a hormone that has long been known to have a large number of physiological functions, especially for the regulation of glucose homeostasis, through the stimulation of its GLP-1R receptor. More recently, it has been shown that the GLP-1 / GLP-1R system is also present at the level of the Central Nervous System where it is involved in memory formation and consolidation, as well as in mechanisms conferring protection to neurons from cell death. However, little is known about the effects that this receptor system has on central neurotransmission. Using immunofluorescence techniques and functional methods, this study showed that GLP-1R receptors are present on synaptic buttons of glutamatergic and GABAergic neurons where they are able to stimulate the exocytotic release of the respective transmitters by activating adenylate cyclase, an effect that is prevented by the selective antagonist of such receptors. The ability to increase glutamate and GABA release by activating GLP-1 presynaptic receptors could, therefore, represent one of the mechanisms by which the neurohormone mediates pro-mnesic and neuroprotective effects that could be exploited for new therapies of neurodegenerative disorders, such as Parkinson’s and Alzheimer’s diseases.
Olivero G, Grilli M, Vergassola M, Bonfiglio T, Padolecchia C, Garrone B, Di Giorgio FP, Tongiani S, Usai C, Marchi M, Pittaluga A. (2018). 5-HT2A-mGlu2/3 receptor complex in rat spinal cord glutamatergic nerve endings: a 5-HT2A to mGlu2/3 signalling to amplify presynaptic mechanism of auto-control of glutamate exocytosis. Neuropharmacology, 133:429-439.
Although the existence and the role of presynaptic-release-regulating mGlu2/3 autoreceptors in CNS is well documented, the new evidence showing that these receptors heterodimerize with non-glutamatergic metabotropic receptors hugely increased the complexity of the scenario. It is the case of 5-H T2Areceptors that were recently reported to associate to mGlu2/3 receptors in the cortex of mammals (Delille et al., 2012, Neuropharmacology 62, 2184). We recently characterized the mGlu2/3 autoreceptors controlling glutamate exocytosis in spinal cord nerve endings. In the present study we extended the research to investigate whether these terminals also possess presynaptic release-regulating 5-HT2A heteroreceptors functionally coupled to mGlu2/3 autoreceptors. Our results unveiled that mGlu2/3 and 5-HT2A receptors colocalize on spinal cord nerve terminals and functional cross-talk in an antagonist-like manner to control glutamate exocytosis. In particular, blockade of 5HT2ARs with selected antagonists increased the mGlu2/3-mediated inhibition of glutamate exocytosis. The gain of function of mGlu2/3Rs relies on an increased expression of the mGlu2/3 receptor protein, consistent with a rapid changes in the number of these autoreceptors in nerve termianls. The observation is particularly intriguing since it indirectly implies i) the existence of a ready-releasable pool of mGlu2/3 receptor proteins in spinal cord glutamatergic nerve endings, ii) the in-out trafficking of mGlu2/3 receptor in terminal plasmamembranes, iii) that antagonists acting at co-localized receptors modulate these rapid in-out movements. Whatever the mechanisms accounting for the changes in the expression of the mGlu2/3 receptor proteins in synaptosomal plasmamembranes, our observations provide the first demonstration that 5-HT2A antagonists act as “indirect positive allosteric modulator” of presynaptic release-regulating mGlu2/3 autoreceptors in rat spinal cord.