1 Huntington’s disease

2 Huntington’s Disease symptoms: An autosomal dominant, heritable disease causing uncontrolled movement of the arms, legs, head, face, and upper trunk. Cognitive impairments in reasoning skills, memory, concentration, judgment, and organization ability Mood changes, especially depression, anxiety, and anger/irritability Frequently, obsessive-compulsive disorder (OCD) develops from alz.org

3 Onset and prognosis of HDNote: juvenile HD is distinct from adult HD in both symptoms & onset! Adult HD: between yrs. Progressive disorder over yrs. Drugs and cell transplants can manage symptoms, but do not change outcome Fasting every other day may improve symptoms & survival (82Q mouse study: PNAS 2003; 100(5): ) Ultimately, patient dies from complications

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5 Early-stage: subtle changes in coordination Stages of HD Early-stage: subtle changes in coordination some involuntary movements (chorea) difficulty thinking about problems depressed or irritable mood overall, less productive at work and home Middle-stage: movement disorder is problematic medication can relieve chorea occupational/physical therapists may be needed for controlling voluntary movemt diminished speech & difficulty swallowing from hdsa.org/what-is-HD

6 Late-stage: total dependence on others for care Stages of HD Late-stage: total dependence on others for care choking is a major concern chorea may be severe or absent inability to walk or speak language comprehension is intact awareness of family & friends is intact death: result of complications, eg. choking or pneumonia, heart failure *All stages of HD: weight loss is a complication from hdsa.org/what-is-HD

7 Huntington’s chorea: caused by mutations in the huntingtin gene (IT-15) located on chromosome 4Huntingtin has a polyglutamine (“polyQ”) repeat sequence encoded by a triplet codon, CAG, which is inherently unstable In Huntington’s disease, the CAG repeat becomes abnormally expanded to extend the number of polyQ repeats Expansion creates a “sticky” protein which aggregates and acts as a molecular sink for interacting factors, generating large intracellular lumps

8 HD is only one of a family of trinucleotide repeat diseasesfrom Wikipedia.org

9 Neurobiological aspects & pathology of HD Progressive neurodegeneration with loss of efferent GABAergic medium spiny neurons in the striatum, and in cortical neurons Animal models include monkey, mouse, fly, fish, and worm HD gene IT-15 discovered in 1993 Pathology of HD (postmortem): described on a scale of 0-4, with 0= 30-40% neuronal loss in head of caudate nucleus; 1= atrophy & neuron loss in the tail of caudate; 2-3= progressive, severe gross atrophy of striatum; 4= most severe HD, with atrophy of striatum & up to 95% neuron loss Zuccato et al, Physiol Rev 2010; 90:

10 Striatal neurons of the basal ganglia are first to die in HD brain

11 Striatal medium-sized spiny neurons (MSNs): 95% of striatuminhibitory GABAergic projection neurons receive input from DA neurons in VTA, SN; Glu neurons in PFC, MCx project to basal ganglia structures uniquely susceptible targets of toxicity in HD

12 Brain regions affected in HD: primary= basal ganglia (STM/GP/TH); cortical neurons affected via BG defects from hdsa.org/what-is-HD

13 The basal ganglia model: Motor cortex  Str  BG  CxThe basis of the model resides in the striatopallidal connections via the direct and the indirect projections (Fig. 5), which have an opposite functional effect on basal ganglia output (Albin et al 1989; DeLong 1990). Activation of GABAergic MSNs that give rise to the direct pathway inhibits GPi/SNr tonic activity, inducing a pause of neuronal firing. At the same time, activation of MSNs of the indirect pathway first inhibits GPe neurons, followed by disinhibition of the STN, which then excites GPi/SNr neurons. Because pauses of neuronal firing in the basal ganglia output are associated with the occurrence of an action (e.g., an eye movement saccade), and discharges are associated with stopping or halting movement, the direct and indirect pathways are viewed as opposite functional projection systems that facilitate and inhibit movements and behaviors, respectively also Cold Spring Harb Perspect Med 2012;2:a009621

14 In early-stage HD, D2-type MSNs are lost first

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16 HD: increased inhibition of STN+ disinhibited thalamus = netHD: increased inhibition of STN+ disinhibited thalamus = net increased movement from web.stanford.edu [HOPES project]

17 HD: disinhibition of striatum + more inhibition from thalamus = slower controlled movementfrom web.stanford.edu [HOPES project]

18 Cellular mechanism for neurodegeneration in HD:trafficking defects endocytosis defects increased mitochondrial fission transcriptional repression

19 Mutant huntingtin binds to HAP1 instead of HIP1endocytosis HAP-1: transport increased # polyQ’s = more HAP1 binding from web.stanford.edu [HOPES project]

20 Mutant Htt increases mitochondrial fission in rat cortical neuronsSong et al, Nat Med Mar; 17(3): 377–382

21 Trafficking defect in Htt-78Q transgenic neuronsasterisk: anterograde diamond: retrograde arrow: fixed point Gunawardena et al, Neuron 2003; 40: 25-40

22 Huntingtin: many posttranslational modifications1st 17 aa’s: amphipathic alpha-helix for mito association (also Golgi/ER); NES/NLS may transport proteins from nucleus  cytoplasm Zuccato et al, Physiol Rev 2010; 90:

23 Functions of Htt’s posttranslational modificationsPhosphorylation (Ser 13/16) of Htt: facilitates vesicular trafficking, clearance of mutant huntingtin, confers neuroprotection (wt Htt) Acetylation of Htt: required for targeting protein to macroautophagy pathway Sumoylation of Htt: modulates localization, activity, and stability inside cell Palmitoylation of Htt: regulates vesicular trafficking

24 Mutant Htt: impairs transcription, axon transport, mito function, & cell survival

25 Therapies for HD: Anti-VMAT (decrease DA-mediated chorea) Histone deacetylase inhibitors Exogenous supply of BDNF Dietary restriction (BDNF & hyperglycemia)

26 Tetrabenazine (TBZ): effective anti-chorea drug

27 Decreased histone deacetylases Sir2 & Sirt2 ameliorate mortality & eye neurodegeneration in Drosophila Htt-Q93 model HDACs: inhibit transcription by decreasing accessibility of DNA to RNA polymerases; in HD, transcription is abnormal Human Mol Gen, 2008, Vol. 17, No –3775

28 Novel approach to stem cell therapy for HDMSCs: do not become neurons or glia; osteoblasts, chondrocytes, myocytes, adipocytes (remote supply of BDNF)

29 Dietary restriction delays HD onset, increases survival, & increases body weightDuan et al, PNAS 􏰀2003 􏰀 vol. 100 􏰀 no. 5 􏰀 2911–2916

30 Dietary restriction: BDNF or autophagy?

31 Alzheimer’s disease

32 Dementia: A chronic disorder of mental processes caused by brain disease or injury, marked by memory disorders, personality changes & impaired reasoning Alzheimer’s disease: A type of progressive dementia that causes problems with memory, thinking & behavior, and is characterized pathologically by neurofibrillary tangles and amyloid plaques *Alzheimer’s: the most common form of dementia (60-80% of all dementia cases) Alzheimer’s Association (alz.org)

33 Common types of dementia:Alzheimer’s disease (50-70%) Vascular dementia (15-30%) Dementia w/ Lewy bodies (DLB; 10-25%) Frontotemporal lobe dementia (rare) Secondary dementias: Normal pressure hydrocephalus (NPH) Parkinson’s dementia (PDD) AIDS-related dementia Alcohol-related dementia

34 Alzheimer’s: not a normal type of age-related memory loss

35 Different types of memory have different brain compartmentslong-term memory storage HI memories: can be consolidated and move to other brain areas emotional memory storage processes implicit memory Left HI: explicit verbal memories Right HI: explicit visual images/locations

36 Diagnosis of Alzheimer’s diseaseMini-mental state evaluation (MMSE): -questionnaire that measures cognitive impairment -examines attention/calculation, recall, language, ability to follow commands, & orientation DSM-IV criteria for AD-type dementia: development of multiple cognitive deficits manifested by memory impairment + one of the following: apraxia, aphasia, agnosia, executive function disturbances Clock-drawing test (CDT): -patient is instructed to draw a clock showing a specific time

37 Neuroimaging to detect signs of Alzheimer’s disease Neuroimaging technology Findings & interpretation PiB PET: fibrillar Aβ imaging w/ 11C-Pittsburgh Compound B (PiB); crosses blood-brain barrier Aβ initially distributed throughout neoCx; spreads to temporal lobe, HI, and deep nuclei FDG-PET: PET scan w/ 18F-fluorodeoxyglucose measures cerebral glucose metabolism/blood flow= indicate synaptic function In AD: decreased metabolism in tempo-parietal Cx at rest= defect in episodic memory fMRI: detects change in neuronal activity during tasks by measuring blood-O2-level-dep. (BOLD) signal In AD, decreased activation of HI during episodic memory tasks DTI: Diffusion tensor imaging detects axon density by measuring diffusion of water in white matter In AD, white matter (fornix, CC) shows decreased density, reflecting neuronal loss

38 Entorhinal cortex & hippocampal atrophy: early signs of ADMRI Cold Spring Harb Perspect Med 2012; 2: a006213

39 Brain regions affected by Alzheimer’s disease

40 Stages of Alzheimer’s diseaseThe course of Alzheimer’s: from subtle changes to inability to communicate early middle late (Alzheimer’s Association)

41 Mild Cognitive ImpairmentDefinition: Cognitive changes that are serious enough to be noticed by the individual or others, but not severe enough to interfere w/ daily life -Individuals w/ MCI have increased risk of developing AD or other dementia (~15-20% people > 65 have MCI) -However, some MCI individuals get better over time -MCI primarily affecting memory= amnestic MCI -MCI affecting thinking skills other than memory= non-amnestic MCI

42 Normal onset vs. early-onset ADCommon form of AD: onset is 60+ years of age Early-onset AD: onset can be in the 30’s Early-onset: due to genetic mutation, usually familial (non-sporadic) Three genes identified cause early-onset AD: Presenilin-1 (PS1) Presenilin-2 (PS2) Amyloid precursor protein (APP) One group of individuals with a very high rate of early-onset Alzheimer’s disease: Down’s syndrome patients Down’s syndrome: trisomy 21 (3x dosage of APP)

43 Alzheimer’s and Down’s syndrome Nearly all Down’s syndrome patients age 40+ have amyloid plaques and neurofibrillary (tau+) tangles in their brains Only a percentage develop AD: ~75% of Down’s patients age 65+ have Alzheimer’s (6-fold higher than average) Early symptoms vary from other AD patients: reduced sociability & enthusiasm, sadness, attention deficit Alzheimer’s Association (alz.org)

44 Intracellular tangles and extracellular plaques in Alzheimer’s brains

45 Impaired ACh release, less choline uptake, receptor deficits in ADSchematic representation of the known and proposed changes in cholinergic neurons that occur in the aged and early AD brain compared with healthy young neurons. Alterations in high-affinity choline uptake, impaired acetylcholine release, deficits in the expression of nicotinic and muscarinic receptors, dysfunctional neurotrophin support (i.e., NGF receptors), and deficits in axonal transport are represented in the early AD neuron either by a decrease in the number of symbols presented or by reduced color intensity. Terry & Buccafusco, J Pharmacol Exper Therap 2003; 306(3):

46 ACh: a major neurotransmitter in the brainACh: functions in both the central & periph NS, at NMJ 7 autonomic NS projections from the basal FB to HI & NCx image: Bruce Blaus

47 Acetylcholine (ACh) AD brain: decreased AChE activity, loss of cholinergic neurons

48 The ACh receptor: a ligand-gated Na+/K+ ion channel2 main AChRs in the brain: α7, α4β2

49 Acetylcholine synaptic transmissionnicotinic AChR: ionotropic muscarinic AChR: metabotropic

50 The amyloid hypothesisMorris et al. Acta Neuropathol Comm 2014, 2:135

51 AICD: intracellular signaling factorAmyloid is cleaved under non-pathogenic conditions by α-secretase and γ-secretase to release AICD & p3 Canter et al, Nature 2016; 539: AICD: intracellular signaling factor

52 Extracellular Aβ aggregates & inhibits synapses to trigger inflammation and cell deathCanter et al, Nature 2016; 539:

53 Amyloid deposition precedes tangles in frontal/temporal lobesThe pathological evolution of Alzheimer’s disease. a | Amyloid plaques and neurofibrillary tangles spread through the brain as the disease progresses. Images are from Spielmeyer’s classic textbook ‘Histopathologie des Nervensystems’ using the Bielschowsky method of silver impregnation to visualize the aggregated proteins that constitute the extracellular plaques and intracellular neurofibrillary tangles172. b | In typical cases of Alzheimer’s disease, amyloid‑β (Aβ) deposition precedes neurofibrillary and neuritic changes with an apparent origin in the frontal and temporal lobes, hippocampus and limbic system (top row). Less commonly, the disease seems to emerge from other regions of the cerebral neocortex (parietal and occipital lobes) with relative sparing of the hippocampus. The neurofibrillary tangles and neuritic degeneration start in the medial temporal lobes and hippocampus, and progressively spread to other areas of the neocortex (bottom row). With the advent of molecular imaging techniques for Aβ and tau, the longitudinal dispersal of pathological changes will become amenable to real-time in vivo study and will not be reliant on post-mortem reconstructions as depicted here. Aβ deposition (stages A, B and C) and neurofibrillary tangles (stages I–VI) are adapted from Braak and Braak17 Masters et al, Nat Rev Dis Prim 2015; Vol 1: 1-18

54 In AD brains, microtubules are destabilized with tangled tau clumps impaired trafficking

55 GSK-3 kinase phosphorylates tau and leads to neuronal tanglesDuan et al. Trans Neurodegen 2012, 1:24

56 Glu neurotransmitter and ADAmyloid beta blocks Glu receptors & transporters, decreasing signaling Excessive synaptic Glu causes NMDAR structural changes/localization and neurotoxicity

57 Normal glutamatergic signaling at the synapseEsposito et al, CNS Neuroscience & Therapeutics 19 (2013) 549–555

58 Aβ-altered glutamate transmission in ADdecreased Ca2+ inhibition of CaMKII/ERK/CREB (& BDNF), activation of survival pathways AND tau phos’n extrasynaptic NR2B NMDARs, decreased synap activ NMDAR blocked LTP *Aβ: blocks EAAT, NR2A+ synaptic NMDARs, & nAChRs net increase in synaptic Glu, decrease in Ca2+ Esposito et al, CNS Neuroscience & Therapeutics 19 (2013) 549–555

59 Neurotrophins and AD NGF BDNF

60 Amyloid interferes with NGF release & transport in ACh neuronsIn AD: decreased ChAT TrkA ChBF neurons

61 Cholinergic BF neurons: decreased levels of NGF in early-stage AD NGF and AD Cholinergic BF neurons: decreased levels of NGF in early-stage AD Increased levels of pro-NGF are present in MCI cerebral cortex *metabolic changes & inflammation alter NGF processing in AD

62 Risk factors vs. deterministic genesRisk factors: increased probability of developing Alzheimer’s, but not a certainty Deterministic genes: mutations in specific genes that will cause Alzheimer’s disease AD risk factors: mild cognitive impairment (MCI) age ApoE4 allele head trauma high blood pressure heart disease

63 Genes with dominant mutations that cause Alzheimer’s: APP, PS-1, PS-2 [deterministic genes] APP -discovered in 1987 -1st gene linked to inherited AD Presenilin-1 -discovered in 1992 -variations in this gene cause the majority of early-onset familial AD Presenilin-2 -discovered in 1993 -3rd gene which causes early-onset AD from alz.org

64 intracranial injection: meningitis heat-killed bacteria controlFig. 1. Ab expression protects against S. Typhimurium meningitis in genetically modified AD mouse models. Transgenic (5XFAD) mice expressing human Ab and mice lacking murine APP (APP-KO) were compared to genetically unmodified littermates [wild type (WT)] for resistance to S. Typhimurium meningitis. One-month-old mice received single ipsilateral intracranial injections of S. Typhimurium, and clinical progression was followed to moribundity. (A to C) Performance of 5XFAD (n = 12) mice compared to WT (n = 11) are shown after infection for survival (P = 0.009) (A), clinical score (P < ) (B), and percent weight loss (P = ) (C). (D) S. Typhimurium load 24 hours after infection in 5XFAD (n = 4) andWT (n = 4) mouse brain hemisphere homogenates shown as mean CFU ± SEM (*P = 0.03 and **P = 0.04). (E) APP-KO mice (n = 15) show a trend (P = 0.104) toward reduced survival compared to WT (n = 15) littermates after infection. (F) No mortality was observed among control sham-infected WT (n = 6) or 5XFAD (n = 6) mice injected with heat-killed S. Typhimurium. Statistical significance was calculated by log-rank (Mantel-Cox) test for survival (A, E, and F), linear regression for clinical score and weight (B and C), and statistical means compared by t test for brain bacterial loads (D). For survival and clinical analysis (A to C), data were pooled from three independent experiments. S.typhimurium intracranial injection: meningitis heat-killed bacteria control Kumar et al, Sci Transl Med 2016; 8(340): 1-16

65 ApoE4 risk factor: reduced amyloid clearance, toxic fragments

66 Effective therapies for AD patients?1) Amyloid immunotherapy or AChE inhibitors: No 2) Exogenous NGF: Maybe 3) Neurostimulation: Promising

67 A4 trial

68 Fibroblasts expressing NGF grafted into AD patient basal forebrain slowed cognitive deficits ~30-40%Tuszynski et al, Nat Med 2005; 11(5):

69 TMS cortical therapy & DBS amygdala-hippocampal therapyCanter et al, Nature 2016; 539:

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