brain research 1497 (2013) 73–84 Available online at www.sciencedirect.com www.elsevier.com/locate/brainres Research Report Pattern of tau hyperphosphorylation and neurotransmitter markers in the brainstem of senescent tau filament forming transgenic mice Kerstin Morcineka,Ã, Christoph Ko¨hlera,Ju¨ rgen Go¨tzb, Hannsjo¨rg Schro¨dera aDepartment of Anatomy II (Neuroanatomy), University of Cologne, Kerpener Strabe 62, 50924 Cologne, Germany bCentre for Ageing Dementia Research (CADR), Queensland Brain Institute (QBI), The University of Queensland, St Lucia Campus (Brisbane), QLD 4072, Australia article info abstract Article history: The early occurrence of brainstem-related symptoms, e.g. gait and balance impairment, Accepted 12 December 2012 apathy and depression in Alzheimer’s disease patients suggests brainstem involvement in Available online 20 December 2012 the initial pathogenesis. To address the question whether tau filament forming mice Keywords: expressing mutated human tau mirror histopathological changes observed in Alzheimer Alzheimer0s disease brainstem, the degree and distribution of neurofibrillary lesions as well as the pattern of Tau hyperphosphorylation cholinergic and monoaminergic neurons were investigated. The expression of the human Neurotransmitter tau transgene was observed in multiple brainstem nuclei, particularly in the magnocellular Brainstem reticular formation, vestibular nuclei, cranial nerve motor nuclei, sensory trigeminal nerve Transgenic mouse nuclei, inferior and superior colliculi, periaqueductal and pontine gray matter, and the red Tauopathy nucleus. Most of the human tau-immunoreactive cell groups also showed tau hyperpho- sphorylation at the epitopes Thr231/Ser235 and Ser202/Thr205, while abnormal tau phosphorylation at the epitope Ser422 or silver stained structures were almost totally lacking. We found no obvious differences in distribution and density of cholinergic and monoaminergic neurons between tau-transgenic and wild type mice. Although numerous brainstem nuclei in our model expressed human tau protein, the development of neurofibrillary tangles, neuropil threads and ghost tangles was rare and likewise its distribution differed largely from Alzheimer0s disease pattern. The number of monoami- nergic neurons remained unchanged in the transgenic mice, while monoaminergic nuclei in Alzheimer brainstem showed a distinct neuronal loss. However, the distribution of Abbreviations (according to Hof et al., 2000): AD, Alzheimer0s disease; AMB, ambiguous nucleus; ChAT, choline acetyltransferase enzyme; DMX, dorsal motor nucleus of the vagus; DR, dorsal nucleus of the raphe; ECU, external cuneate nucleus; GRN, gigantocellular reticular nucleus; IC, inferior colliculus; III, oculomotor nucleus; ir/ IR, immunoreactive/ immunoreactivity; IRN, intermediate reticular nucleus; ISN, inferior salivatory nucleus; IV, trochlear nucleus; LC, locus coeruleus; LRN, lateral reticular nucleus; LVN, lateral vestibular nucleus; MVN, medial vestibular nucleus; NFTs, neurofibrillary tangles; PAG, periaqueductal gray matter; PB, parabrachial nucleus; PG, pontine gray matter; PGRNl, lateral paragigantocellular nucleus; PPN, pedunculopontine nucleus; PRN, pontine reticular nucleus; PSP, progressive supranuclear palsy; RN, red nucleus; RVLN, rostroventrolateral reticular nucleus; SC, superior colliculus; SPV, spinal trigeminal nucleus; SVN, superior vestibular nucleus; TH, tyrosine hydroxylase; V, motor trigeminal nucleus; VII, facial nucleus; VLL, ventral nucleus of the lateral lemniscus; XII, hypoglossal nucleus ÃCorresponding author. Fax: þ49 221 478 5318. E-mail address: [email protected] (K. Morcinek). 0006-8993/$ - see front matter & 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.brainres.2012.12.016 74 brain research 1497 (2013) 73–84 pretangle-affected neurons in the tau-transgenic mice partly resembled those seen in progressive supranuclear palsy, presenting these animals as a model to examine brainstem pathogenesis of progressive supranuclear palsy. & 2013 Elsevier B.V. All rights reserved. 1. Introduction Procter et al., 1988). The number of serotonergic neurons in the DR - source of the ascending serotonergic transmitter system Abnormally phosphorylated forms of the microtubule asso- to numerous cortical and subcortical regions of the forebrain ciated protein tau - physiologically abundantly expressed in (Ru¨ betal.,2000) - declines to 50% of the control value in AD neurons and glial cells (Lee et al., 2001) - constitute the major subjects (Zweig et al., 1988). Dysfunction within this system component of neurofibrillary tangles (NFTs) and neuropil can cause disrupted sleeping pattern, depressive mood swings, threads in the brain tissue of patients suffering from and inadequate affective control (Ru¨ betal.,2000). Whether Alzheimer0s disease (AD) (Mercken et al., 1992). In addition to these deficits in neurotransmission are linked to the occur- telencephalic changes (Braak and Braak, 1991a) evidence has rence of neurofibrillary lesions in cholinergic and monoami- been presented for the early appearance of tau pathology in nergic nuclei, e.g. in the noradrenergic locus coeruleus or the brainstem nuclei of AD subjects (Simic et al., 2009). In the serotonergic oral raphe complex, is not finally clarified. midbrain, the oral raphe nuclei, i.e. central linear, central Notwithstanding the findings of NFTs and neuropil threads in superior and dorsal raphe nucleus (DR) show tau deposition brainstem nuclei and the neuronal loss in monoaminergic nuclei in correlation with the staging system of Braak and Braak (Ru¨ b in the reticular formation, no animal model reflecting brainstem et al., 2000). During the disease process the supratrochlear changes that are observed in AD patients has been described yet. subunit of the DR is already affected by tangle formation prior Only sparse information about neurofibrillary lesions in the to the transentorhinal stage (Grinberg et al., 2009). The peria- brainstem of tau transgenic mice is obtainable (Delobel et al., queductal gray matter (PAG), the pedunculopontine nucleus 2008; Dutschmann et al., 2010; Menuet et al., 2011; Overk et al., (PPN), the parabrachial nucleus (PB) (German et al., 1987; Parvizi 2009; Probst et al., 2000). Therefore the objective of the present et al., 2001) and the locus coeruleus (LC) (Hirano and study was to investigate the degree and localization of neurofi- Zimmerman, 1962; Ishii, 1966; Parvizi et al., 2001)–inclusive brillary lesions as well as the distribution of cholinergic, cate- the oral raphe nuclei constituting the main components of the cholaminergic and serotonergic neurons throughout the brainstem ascending arousal system – exhibit insoluble fila- brainstem of senescent P301L tau transgenic pR5 mice. mentous tau during isocortical stages. Within the reticular formation – containing several control centers of vitally impor- tant functions, e.g. cardiovascular, respiratory and visceral 2. Results regulation - the intermediate reticular nucleus (IRN) develops NFTs (Parvizi et al., 2001; Ru¨ betal.,2001). In particular the 2.1. Location of anti-tau antibodies in the brainstem recent finding of intraneuronal pretangle material in the LC of of P301L tau transgenic pR5 mice children and young adults may indicate that the pathologic process leading to abnormal tau pathology begins in selected 2.1.1. HT7-immunoreactivity (IR) subcorticalnuclei(Braak and Del Tredici, 2011b). The phosphorylation-independent anti-tau antibody HT7 Along with tau pathology, several neurotransmitter systems specifically recognizing the human tau protein was used to in AD brains are affected by dysfunction, loss of neurons and detect neurons expressing the transgene construct. reduced transmitter concentrations (Palmer and DeKosky, In the reticular formation from caudal to rostral a moderate 1993). Particularly the cholinergic basal nucleus of Meynert number of HT7-positive neurons was observed in the lateral and septal nuclei with projections to almost all cortical areas, reticular nucleus (LRN) and GRN (Fig. 1a), fewer neurons were the hippocampus and the thalamus are severely impaired in a stained in the IRN (Fig. 1b), lateral part of paragigantocellular subset of AD brains (Herholz, 2008; McGeer et al., 1984). The (PGRNl), parvocellular, mesencephalic and pontine (PRN) loss and functional disturbance of cholinergic forebrain neu- reticular nucleus (Fig. 2, Bregma À7.5 mm to À4.4 mm). In rons is associated with a reduction of cholinergic neurotrans- two tau transgenic mice a solitary HT7-labeled perikaryon mission, choline acetyltransferase and acetylcholinesterase was located in the rostroventrolateral reticular nucleus enzyme activity (Francis et al., 1999; Herholz, 2008; Procter (RVLN). No expression of human tau was detected in nuclei et al., 1988). Likewise, the content of the catecholamine of the raphe in the median region and in the LC and PB in the noradrenaline is reduced in AD temporal and frontal cortices lateral region of the formatio reticularis. The nucleus raphe (Palmer and DeKosky, 1993). The neuron density in the pontine obscurus showed a single labeled neuron in two individuals. LC – major nucleus of origin of corticopetal noradrenergic In the caudal medulla oblongata one to five human tau- fibers - is significantly diminished (Zweig et al., 1988). Also positive neurons per section were found in the external the concentration of the neurotransmitter serotonin and the cuneate nucleus (ECU) and in the more medially located number