The purpose of these experiments was to determine the effects of cholinergic depletion on spontaneous and evoked activity of neurons in the different layers of the posteromedial barrel subfield (PMBSF) of the rat somatosensory cortex. Acetylcholine neurons in nucleus basalis of Meynert (NBM) were selectively lesioned with an immunotoxin (IT), 192 IgG-saporin. Spontaneous activity was significantly lower in layers II-III, Va, and VI in IT-injected animals compared to control animals. Evoked activity was significantly lower in layers II-III, IV, Vb, and VI of IT-injected animals compared to control animals. The largest difference was observed in layer Vb. Thus, cholinergic depletion causes significant changes in the magnitude of spontaneous and evoked activity but these differences are not completely in register with one another.

abstract: Alzheimer’s disease (AD) is characterized by the degeneration of cholinergic basal forebrain (CBF) neurons in the nucleus basalis of Meynert (nbM), which provides the majority of cholinergic input to the cortical mantle and together form the basocortical cholinergic system. Histone deacetylase (HDAC) dysregulation in the temporal lobe has been associated with neuronal degeneration during AD progression. However, whether HDAC alterations play a role in cortical and cortically-projecting cholinergic nbM neuronal degeneration during AD onset is unknown. In an effort to characterize alterations in the basocortical epigenome semi-quantitative western blotting and immunohistochemistry were utilized to evaluate HDAC and sirtuin (SIRT) levels in individuals that died with a premortem clinical diagnosis of no cognitive impairment (NCI), mild cognitive impairment (MCI), mild/moderate AD (mAD), or severe AD (sAD). In the frontal cortex, immunoblots revealed significant increases in HDAC1 and HDAC3 in MCI and mAD, followed by a decrease in sAD. Cortical HDAC2 levels remained stable across clinical groups. HDAC4 was significantly increased in prodromal and mild AD compared to aged cognitively normal controls. HDAC6 significantly increased during disease progression, while SIRT1 decreased in MCI, mAD, and sAD compared to controls. Basal forebrain levels of HDAC1, 3, 4, 6 and SIRT1 were stable across disease progression, while HDAC2 levels were significantly decreased in sAD. Quantitative immunohistochemistry was used to identify HDAC2 protein levels in individual cholinergic nbM nuclei immunoreactive for the early phosphorylated tau marker AT8, the late-stage apoptotic tau marker TauC3, and Thioflavin-S, a marker of mature neurofibrillary tangles (NFTs). HDAC2 nuclear immunoreactivity was reduced in individual cholinergic nbM neurons across disease stages, and was exacerbated in tangle-bearing cholinergic nbM neurons. HDAC2 nuclear reactivity correlated with multiple cognitive domains and with NFT formation. These findings identify global HDAC and SIRT alterations in the cortex while HDAC2 dysregulation contributes to cholinergic nbM neuronal dysfunction and NFT pathology during the progression of AD. / Dissertation/Thesis / Doctoral Dissertation Neuroscience 2018

Alzheimer’s disease (AD) is a degenerative disorder where the distribution of pathology throughout the brain is not random but follows a predictive pattern used for pathological staging. While the involvement of defined functional systems is fairly well established for more advanced stages, the initial sites of degeneration are still ill defined. The prevailing concept suggests an origin within the transentorhinal and entorhinal cortex (EC) from where pathology spreads to other areas. Still, this concept has been challenged recently suggesting a potential origin of degeneration in nonthalamic subcortical nuclei giving rise to cortical innervation such as locus coeruleus (LC) and nucleus basalis of Meynert (NbM). To contribute to the identification of the early site of degeneration, here, we address the question whether cortical or subcortical degeneration occurs more early and develops more quickly during progression of AD. To this end, we stereologically assesses neurone counts in the NbM, LC and EC layer-II in the same AD patients ranging from preclinical stages to severe dementia. In all three areas, neurone loss becomes detectable already at preclinical stages and is clearly manifest at prodromal AD/MCI. At more advanced AD, cell loss is most pronounced in the NbM > LC > layer-II EC. During early AD, however, the extent of cell loss is fairly balanced between all three areas without clear indications for a preference of one area. We can thus not rule out that there is more than one way of spreading from its site of origin or that degeneration even occurs independently at several sites in parallel.

Η αντιπονεκτίνη και οι υποδοχείς αντιπονεκτίνης, AdipoR1 και AdipoR2, αποτελούν συστατικά στοιχεία των ενεργειακών ομοιοστατικών μηχανισμών στους περιφερικούς ιστούς. Σύμφωνα με πρόσφατες μελέτες, η αντιπονεκτίνη φαίνεται, επιδρώντας σε κεντρικά νευρωνικά κυκλώματα, να συμμετέχει στη ρύθμισης πρόσληψης τροφής και κατανάλωσης ενέργειας. Σκοπός της παρούσας μελέτης ήταν η διερεύνηση της πιθανής έκφρασης και της κατανομής της αντιπονεκτίνης και των υποδοχέων της στην ανθρώπινη υπόφυση, στον υποθάλαμο και σε άλλες περιοχές του ανθρώπινου εγκεφάλου. Τομές υπόφυσης, υποθαλάμου και της παρακείμενης βασικής τηλεγκεφαλικής περιοχής, εγκεφαλικού φλοιού και παρεγκεφαλίδας μονιμοποιημένες σε ουδέτερη φορμόλη και εγκλεισμένες σε παραφίνη, από σαράντα περιστατικά, μελετήθηκαν ιστολογικά με τη χρήση ηωσίνης-αιματοξυλίνης, και των ειδικών χρώσεων PAS-orange G και luxol fast blue-cresyl violet. Εν συνεχεία, εφαρμόσθηκε απλή και διπλή ανοσοϊστοχημική μέθοδος, χρησιμοποιώντας ειδικά αντισώματα έναντι της αντιπονεκτίνης, του AdipoR1 και AdipoR2, της ακετυλομεταφοράσης της χολίνης, της FSH, LH, TSH, GH, ACTH και προλακτίνης. Ο μέσος όρος (± SD) ηλικίας και δείκτη μάζας σώματος (ΒΜΙ) των υπό εξέταση περιπτώσεων ήταν 56 (±18) έτη και 27 (±5) kg/m2, αντίστοιχα. Έντονη έκφραση της αντιπονεκτίνης παρατηρήθηκε στον πρόσθιο λοβό (pars distalis/PD) της υπόφυσης και στο χοανικό δακτύλιο (pars tuberalis/PT). Ειδικότερα, ισχυρή ανοσοϊστοχημική χρώση για την αντιπονεκτίνη παρατηρήθηκε στα κύτταρα που παράγουν GH, FSH, LH , TSH και FSH, LH, TSH, στον πρόσθιο λοβό και στο χοανικό δακτύλιο αντίστοιχα.. Στο PD, ισχυρή έως μέτρια έκφραση του AdipoR1 και AdipoR2 ανιχνεύθηκε στους ίδιους κυτταρικούς τύπους στους οποίους εντοπίσθηκε και η αντιπονεκτίνη. Δεν παρατηρήθηκε ανοσοθετικότητα για τους υποδοχείς της αντιπονεκτίνης στα κύτταρα του ΡT. Έντονη ανοσοϊστοχημική χρώση για τον AdipoR1 παρουσίασαν οι νευρώνες της πλάγιας υποθαλαμικής περιοχής και του βασικού πυρήνα του Meynert (NBM). Η έκφραση της αντιπονεκτίνης και των υποδοχέων της στην ανθρώπινη υπόφυση ενδεχομένως αποτελεί μία ένδειξη της ύπαρξης ενός τοπικού ρυθμιστικού συστήματος, το οποίο ασκεί τροποποιητικές δράσεις στους ενδοκρινικούς άξονες. Επιπρόσθετα, η παρουσία του AdipoR1 στον υποθάλαμο και στο NBM υποδεικνύει ότι η αντιπονεκτίνη μπορεί να 118 συμμετέχει σε κεντρικά νευρωνικά σηματοδοτικά μονοπάτια, ελέγχοντας την ενεργειακή ομοιόσταση και άλλες εγκεφαλικές λειτουργίες. / Adiponectin and its receptors, AdipoR1 and AdipoR2, constitute integral components of energy homeostatic mechanism, in peripheral tissues. Recent studies have implicated adiponectin in central neural networks regulating food intake and energy expenditure. The present study aimed at investigating the possible expression and distribution of adiponectin and its receptors in human pituitary gland, hypothalamus and different brain areas. Sections of the pituitary gland, hypothalamus and adjacent basal forebrain area, cerebrum and cerebellum from forty autopsy cases, were examined using H&E, PAS-Orange G, luxol fast blue/cresyl violet stains and single and double immunohistochemistry using adiponectin, AdipoR1, AdipoR2, choline acetyltransferase, FSH, LH, TSH, GH, ACTH and prolactinspecific antibodies. Age and BMI mean values ± SD of the autopsy cases were 56±18 years and 27±5 kg/m2, respectively. Strong adiponectin expression was observed in pituitary gland. In pars distalis (PD), adiponectin localized in GH, FSH, LH and TSH-producing cells and in pars tuberalis (PT) in FSH, LH and TSH-producing cells. Strong to moderate expression of AdipoR1 and AdipoR2 was observed in PD by the same cell types as adiponectin. No immunoreactivity for adiponectin receptors was noted in cells of PT. Intense AdipoR1 immunostaining was observed in neurons of lateral hypothalamic area and of nucleus basalis of Meynert (NBM). Adiponectin and its receptors expression in human pituitary might indicate the existence of a local system, modulating endocrine axes. Furthermore, the presence of AdipoR1 in hypothalamus and NBM suggests that adiponectin may participate in central neural signaling pathways controlling energy homeostasis and higher brain functions.

Alzheimer’s disease (AD) is a degenerative disorder where the distribution of pathology throughout the brain is not random but follows a predictive pattern used for pathological staging. While the involvement of defined functional systems is fairly well established for more advanced stages, the initial sites of degeneration are still ill defined. The prevailing concept suggests an origin within the transentorhinal and entorhinal cortex (EC) from where pathology spreads to other areas. Still, this concept has been challenged recently suggesting a potential origin of degeneration in nonthalamic subcortical nuclei giving rise to cortical innervation such as locus coeruleus (LC) and nucleus basalis of Meynert (NbM). To contribute to the identification of the early site of degeneration, here, we address the question whether cortical or subcortical degeneration occurs more early and develops more quickly during progression of AD. To this end, we stereologically assesses neurone counts in the NbM, LC and EC layer-II in the same AD patients ranging from preclinical stages to severe dementia. In all three areas, neurone loss becomes detectable already at preclinical stages and is clearly manifest at prodromal AD/MCI. At more advanced AD, cell loss is most pronounced in the NbM > LC > layer-II EC. During early AD, however, the extent of cell loss is fairly balanced between all three areas without clear indications for a preference of one area. We can thus not rule out that there is more than one way of spreading from its site of origin or that degeneration even occurs independently at several sites in parallel.