Regionally selective changes in brain lysosomes occur in the transition from young adulthood to middle age in rats

doi:10.1016/S0306-4522(00)00021-X

Neuroscience

Volume 97, Issue 2, April 2000, Pages 395-404

https://doi.org/10.1016/S0306-4522(00)00021-X Get rights and content

Abstract

The possibility that brain aging in rats exhibits regional variations of the type found in humans was studied using lysosomal chemistry as a marker. Age-related (two vs 12 months; male Sprague–Dawley) differences in cathepsin D immunostaining were pronounced in the superficial layers of entorhinal cortex and in hippocampal field CA1, but not in neocortex and field CA3. Three changes were recorded: an increase in the intraneuronal area occupied by labeled lysosomes; clumping of immunopositive material within neurons; more intense cytoplasmic staining. Western blot analyses indicated that the increases involved the active forms of cathepsin D rather than their proenzyme. Shrinkage of cathepsin-D-positive neuronal cell bodies was observed in entorhinal cortex but not in neocortical sampling zones. Age-related lysosomal changes as seen with cathepsin B immunocytochemistry were considerably more subtle than those obtained with cathepsin D antibodies. In contrast, a set of glial and/or vascular elements located in a distal dendritic field of the middle-aged hippocampus was much more immunoreactive for cathepsin B than cathepsin D. The areas exhibiting sizeable changes in the present study are reported to be particularly vulnerable to aging in humans.

The results thus suggest that aspects of brain aging common to mammals help shape neurosenescence patterns in humans.

Section snippets

Experimental procedures

All animal experiments were carried out in accordance with the National Institute of Health guide for the care and use of laboratory animals, and all efforts were made to minimize animal suffering.

Age-related changes in regional levels of cathepsin D

A section processed for cathepsin D immunoreactivity (ir) in the cortical telencephalon of a two-month-old rat is shown in Fig. 1. Labeling was discrete and restricted to cell bodies; with few exceptions, it did not differ across subdivisions to a degree greater than would be expected from variations in cell packing densities. Neurons in cortical layer V (* in Fig. 1A, B), the hilus of the dentate gyrus, and hippocampal pyramidal neurons were more densely stained than cells in the surrounding

Discussion

These results demonstrate that pronounced changes in lysosomes and certain hydrolases develop by middle age in select areas of rat telencephalon. The affected zones correspond to areas of human brain that are particularly vulnerable to aging and Alzheimer's disease. Accordingly, regionally differentiated neurosenescence in humans10., 11., 15., 43. could, in part, be the outcome of a general feature of mammalian brain aging.

The changes in lysosomal size and location in the 12-month animals are

Acknowledgments

The authors thank David Ko, Zain Vally-Mahomed, and Jordan Tran for laboratory assistance. This work is supported by a grant from the National Institute of Aging (AG00538) to G.L.

References (58)

X. Bi et al.
Lysosomal protease inhibitors cause meganeurite formation and phosphorylated tau abnormalities in entorhinal–hippocampal regions vulnerable to Alzheimer's disease
Expl Neurol.
(1999)
M.M. Bradford
A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein–dye binding
Analyt. Biochem.
(1976)
N. Chevallier et al.
Cathepsin D displays in vitro beta-secretase-like specificity
Brain Res.
(1997)
L.B. Creemers et al.
Participation of intracellular cysteine proteinases, in particular cathepsin B, in degradation of collagen in periosteal tissue explants
Matrix Biol.
(1998)
M. Dezawa et al.
Glial cells in degenerating and regenerating optic nerve of the adult rat
Brain Res. Bull.
(1999)
K.B. Hoffman et al.
Beta-amyloid increases cathepsin D levels in hippocampus
Neurosci. Lett.
(1998)
W.G. Honer et al.
Regional synaptic pathology in Alzheimer's disease
Neurobiol. Aging
(1992)
H. Jung et al.
Age-related changes in ultrastructural features of cathepsin B- and D-containing neurons in rat cerebral cortex
Brain Res.
(1999)
P. Landfield et al.
Hippocampal aging in rats: a morphometric study of multiple variables in semithin sections
Neurobiol. Aging
(1981)
C.T. Moore et al.
Synaptic density in the arcuate nucleus of female rats approaching middle age
Neurosci. Lett.
(1998)

Y. Nakamura et al.

Lysosome instability in aged rat brain

Neurosci. Lett.

(1989)

H. Nakanishi et al.

Age-related changes in activities and localizations of cathepsin D, E, B, and L in the rat brain tissues

Expl Neurol.

(1994)

D.P. Purpura et al.

Fine structure of meganeurites and secondary growth processes in feline GM1-gangliosidosis

Brain Res.

(1978)

D.P. Purpura et al.

Distortion of neuronal geometry and formation of aberrant synapses in neuronal storage disease

Brain Res.

(1976)

M.M. Shores et al.

Tyrosine hydroxylase mRNA is increased in old age and norepinephrine uptake transporter mRNA is decreased in middle age in locus coeruleus of Brown-Norway rats

Brain Res.

(1999)

S.U. Walkley et al.

Distribution of ectopic neurite growth and other geometrical distortions of CNS neurons in feline GM2 gangliosidosis

Brain Res.

(1990)

A.P. Yong et al.

Lysosomal dysfunction results in lamina specific meganeurite formation in frontal cortex

Expl Neurol.

(1999)

B.M. Austen et al.

Cleavage of a beta-amyloid precursor sequence by cathepsin D

Biomed. Peptides Proteins Nucleic Acids

(1995)

B. Bahr et al.

Induction of β-amyloid containing polypeptides in hippocampus: evidence for a concomitant loss of synaptic proteins and interactions with an excitotoxin

Expl Neurol.

(1994)

M. Bannay-Schwartz et al.

The distribution of cathepsin D activity in adult and aging human brain regions

J. Neurochem.

(1992)

E. Bednarski et al.

Cytosolic proteolysis of tau by cathepsin D in hippocampus following suppression of cathepsins B and L

J. Neurochem.

(1996)

E. Bednarski et al.

Selective suppression of cathepsin L results from elevations in lysosomal pH and is followed by proteolysis of tau protein

NeuroReport

(1998)

E. Bednarski et al.

Suppression of cathepsins B and L causes a proliferation of lysosomes and the formation of meganeurites in hippocampus

J. Neurosci.

(1997)

N. Bodeutsch et al.

Unilateral injury to the adult rat optic nerve causes multiple cellular responses in the contralateral site

J. Neurobiol.

(1999)

Braak H. (1984) Architectonics as seen by lipofuscin stains. In: Cerebral Cortex (eds Peters A. and Jones E.G.), pp....

H. Braak et al.

Neuropathological staging of Alzheimer-related changes

Acta neuropath.

(1991)

Braak H. and Braak E. (1994) Pathology of Alzheimer's disease. In Neurodegenerative Diseases (ed. Calne D. B.), pp....

H. Braak et al.

Pigmentoarchitectonic pathology of the isocortex in juvenile neuronal ceroid-lipofuscinosis: axonal enlargements in layer IIIab and cell loss in layer V

Acta neuropath.

(1979)

A. Cai et al.

Transplantation of fetal suprachiasmatic nuclei into middle-aged rats restores diurnal Fos expression in host

Am. J. Physiol.

(1997)

Cited by (31)

Lysosomal cathepsins and their regulation in aging and neurodegeneration
2016, Ageing Research Reviews
Citation Excerpt :
Moreover, in human tissue, cathepsin D was found to be present not only in multiple neurons, but also in some glial cells with increasing levels during neuroontogenesis (Dorn et al., 1986). Regionally selective changes in brain lysosomes were observed in the transition from young adulthood to middle age in rats (Bi et al., 2000). Pronounced age-related changes that involved primarily translocation of cathepsin D in neurons from lysosomes to cytosolic granules, were observed also in rat cerebral cortex in more than 35% of cells.
Lysosomes and lysosomal hydrolases, including the cathepsins, have been shown to change their properties with aging brain a long time ago, although their function was not really understood. The first biochemical and clinical studies were followed by a major expansion in the last 20 years with the development of animal disease models and new approaches leading to a major advancement of understanding of the role of physiological and degenerative processes in the brain at the molecular level. This includes the understanding of the major role of autophagy and the cathepsins in a number of diseases, including its critical role in the neuronal ceroid lipofuscinosis. Similarly, cathepsins and some other lysosomal proteases were shown to have important roles in processing and/or degradation of several important neuronal proteins, thereby having either neuroprotective or harmful roles. In this review, we discuss lysosomal cathepsins and their regulation with the focus on cysteine cathepsins and their endogenous inhibitors, as well as their role in several neurodegenerative diseases.
Reconsider Alzheimer's disease by the 'calpain-cathepsin hypothesis'-A perspective review
2013, Progress in Neurobiology
Citation Excerpt :
In the AD brain, cathepsin D immunoreactivity was often localized extracellularly to the senile plaques (Cataldo and Nixon, 1990; Cataldo et al., 1995, 1996). In the rat brain, cathepsin D showed an age-dependent significant increase (Bi et al., 2000, 2003), while cathepsin L showed an age-related remarkable reduction (Nakanishi et al., 1994). A similar increase in the cathepsin D immunoreactivity associated with aging was found in neurons of dogs (Lynch and Bi, 2003).
Alzheimer's disease (AD) is characterized by slowly progressive neuronal death, but its molecular cascade remains elusive for over 100 years. Since accumulation of autophagic vacuoles (also called granulo-vacuolar degenerations) represents one of the pathologic hallmarks of degenerating neurons in AD, a causative connection between autophagy failure and neuronal death should be present. The aim of this perspective review is at considering such underlying mechanism of AD that age-dependent oxidative stresses may affect the autophagic-lysosomal system via carbonylation and cleavage of heat-shock protein 70.1 (Hsp70.1). AD brains exhibit gradual but continual ischemic insults that cause perturbed Ca²⁺ homeostasis, calpain activation, amyloid β deposition, and oxidative stresses. Membrane lipids such as linoleic and arachidonic acids are vulnerable to the cumulative oxidative stresses, generating a toxic peroxidation product ‘hydroxynonenal’ that can carbonylate Hsp70.1. Recent data advocate for dual roles of Hsp70.1 as a molecular chaperone for damaged proteins and a guardian of lysosomal integrity. Accordingly, impairments of lysosomal autophagy and stabilization may be driven by the calpain-mediated cleavage of carbonylated Hsp70.1, and this causes lysosomal permeabilization and/or rupture with the resultant release of the cell degradation enzyme, cathepsins (calpain–cathepsin hypothesis). Here, the author discusses three topics; (1) how age-related decrease in lysosomal and autophagic activities has a causal connection to programmed neuronal necrosis in sporadic AD, (2) how genetic factors such as apolipoprotein E and presenilin 1 can facilitate lysosomal destabilization in the sequential molecular events, and (3) whether a single cascade can simultaneously account for implications of all players previously reported. In conclusion, Alzheimer neuronal death conceivably occurs by the similar ‘calpain-hydroxynonenal-Hsp70.1-cathepsin cascade’ with ischemic neuronal death. Blockade of calpain and/or extra-lysosomal cathepsins as well as scavenging of hydroxynonenal would become effective AD therapeutic approaches.
Upregulation of cathepsin S in the aging and pathological nervous system of mice
2008, Brain Research
Citation Excerpt :
With respect to the phenotype of these CATS expressing cells in the aged brain, we noted that mostly glial cells are concerned. An upregulation of protein expression during aging in glial cells has also been reported for CATB, CATX and CATD (Amano et al., 1995; Bi et al., 2000; Wendt et al., 2007). A numerical increase of glial cells accompanying an increased inflammatory activity has been well described to occur during normal brain aging (Ogura et al., 1994; Sloane et al., 1999; Streit et al., 2004; Stichel and Lübbert, 2007).
Cathepsins have long been regarded enzymes that are primarily involved in general protein turnover within lysosomes. However, more recently, their differential cell and tissue distributions suggest that at least some of them participate in specific cellular processes. Cathepsin S (CATS) is mainly expressed in cells of mononuclear phagocytotic origin and plays a major role in the MHC-II-mediated antigen presentation. Although a central role for CATS in brain function has also been suggested, its localization and regulation in the central nervous system are still poorly understood. In the present study we investigated the regional and cellular expression of CATS in normal, aging and pathological mouse brain. Our studies show that CATS is expressed throughout the adult mouse brain, in particular in microglial cells. In aged mice, CATS protein expression increases in these cells. In addition, it became apparent that in old mice a larger number of neuronal cells stained positive for this protease. At the subcellular level, CATS immunostaining accumulated in granules, indicating a lysosomal localization. In a transgenic mouse model of amyotrophic lateral sclerosis expressing mutant superoxide dismutase 1 (SOD1), CATS transcript and protein levels were significantly upregulated in spinal cord and lower brain regions displaying neuronal degeneration. The majority of strongly immunopositive cells in these regions exhibited microglial morphology. These results suggest that CATS participates in inflammatory processes accompanying aging and pathologies of the CNS.
Exploiting endogenous anti-apoptotic proteins for novel therapeutic strategies in cerebral ischemia
2008, Progress in Neurobiology
Citation Excerpt :
In the brain, potential substrates for μ-calpain include a number of cytoskeletal proteins such as actin-binding proteins (for example, spectrin, actinin, gelsolin, gephyrin, ankyrin, talin, tubulin, microtubules-associated proteins, and neurofilaments [reviewed in Rami, 2003]). Potential substrates for μ-calpain are also membrane proteins such as growth factors receptors and calcium channels, adhesion molecules, ion transporters, enzymes, such as kinases, phosphatases (calcineurin), and phospholipases, cytokines, and transcription factors (Saido et al., 1994; Dosemeci and Reese, 1995; Bednarski et al., 1995; Blomgren et al., 1997; Bi et al., 2000). The strongest evidence for the direct participation of μ-calpain in neurodegeneration has been found upon acute brain ischemia and was highlighted by the cytoprotective effect of calpain inhibitors (Rami and Krieglstein, 1993a,b; Rami and Krieglstein, 1994).
The acute neuronal degeneration in the ischemic core upon stroke is followed by a second wave of cell demise in the ischemic penumbra and neuroanatomically connected sites. This temporally delayed deleterious event of programmed cell death (‘secondary degeneration’) often exceeds the initial damage of stroke and, thus, contributes pivotally to significant losses in neurological functions. In fact, it is the injured neurons in these regions around the ischemic core zone that neuropharmacological prevention is targeting to preserve. Clinical and pre-clinical studies have focussed on neuroprotective interventions with caspase inhibitors, but it remains ambiguous whether diminishing or even silencing these aspartate-specific cysteine proteases are in sum beneficial for the clinical outcome. It is often ignored that caspase inhibitors are able to antagonize calpain and cathepsins, thereby protecting the cytoskeleton from damage. Moreover, there is a point of no return, beyond which interfering with caspases cannot rescue the cell, but spoil the obligate and necessary suicide program such that the cellular environment suffers from by-products of necrosis and secondary inflammation. Here we discuss novel alternative strategies to abrogate the death cascade at the level of the genomic response (transcription factors, NF-κB, CREB, ICER, HIF), of mitochondrial effectors (cytochrome c, Bcl-2, Smac/DIABLO, HtrA2), and of inhibitor of apoptosis proteins (IAPs). IAPs are the only known endogenous proteins that inhibit specifically and with high affinity the activity of both initiator and effector caspases. Based on compelling biochemical evidence, we argue that patronizing the neuronal endogenous anti-apoptotic machinery could be superior to the pharmacological inhibition of caspases at various levels, with regard to specificity, side effects, and the ‘therapeutic window of opportunity’.
Differential expression of cathepsin X in aging and pathological central nervous system of mice
2007, Experimental Neurology
Increasing evidence of a fundamental influence of cathepsins on inflammation has drawn interest in a thorough understanding of their role in physiological and pathological processes. Even though the number of identified cathepsins has more than doubled in the last years, information about their expression, regulation and function in the brain is still incomplete.
In the present study we analyzed the regional, cellular and subcellular localization and the activity of the recently discovered cathepsin X in the normal, developing and pathological mouse brain. Our results show that CATX is: (i) is expressed in almost all cells in the mouse brain with a preference for glial cells; (ii) already widely expressed early in development and age-dependently upregulated in amount and activity; (iii) prominently localized in the lysosomal system but also scattered in the somal cytoplasm in the aged brain; (iv) upregulated in numerous glial cells of degenerating brain regions in a transgenic mouse model of amyotrophic lateral sclerosis; and (v) associated with plaques in a transgenic mouse model and in Alzheimer patients.
These results strongly suggest that cathepsin X is an important player in degenerative processes during normal aging and in pathological conditions.
Calcium-dependent and aspartyl proteases in neurodegeneration and ageing in C. elegans
2003, Ageing Research Reviews
Proteolytic mechanisms have been implicated in the process of ageing, and in many neurodegenerative disorders such as Alzheimer’s, Parkinson’s and Huntington’s diseases, which are most prevalent in old age. Simple model organisms such as the nematode Caenorhabditis elegans and the fruit fly Drosophila melanogaster, which offer the prowess of sophisticated genetic approaches, have contributed to our understanding of ageing and neurodegeneration. Intensive research in these systems has resulted in detailed models of the ageing process, and also of several neurodegenerative diseases, which recapitulate same aspects of the human pathologies. Inappropriate cell death is a major component of these and other devastating conditions such as stroke. The dissection of the molecular mechanisms underlying the process of cell degeneration in ageing is of outmost importance. Evidence from investigations in C. elegans implicates deregulated proteolysis as one major determinant of cellular destruction in neurodegeneration and ageing, and suggests that the process depends critically on the activation of calcium-dependent, calpain proteases and lysosomal aspartyl proteases. Apart from shedding light on important but inadequately understood facets of such phenomena, these discoveries hold promise for developing novel, effective intervention strategies aiming to ameliorate or even counter inappropriate cell death.

View all citing articles on Scopus

View full text

Regionally selective changes in brain lysosomes occur in the transition from young adulthood to middle age in rats

Abstract

Section snippets

Experimental procedures

Age-related changes in regional levels of cathepsin D

Discussion

Acknowledgments

Expl Neurol.

Analyt. Biochem.

Brain Res.

Matrix Biol.

Brain Res. Bull.

Neurosci. Lett.

Neurobiol. Aging

Brain Res.

Neurobiol. Aging

Neurosci. Lett.

Neurosci. Lett.

Expl Neurol.

Brain Res.

Brain Res.

Brain Res.

Brain Res.

Expl Neurol.

Cleavage of a beta-amyloid precursor sequence by cathepsin D

Biomed. Peptides Proteins Nucleic Acids

Induction of β-amyloid containing polypeptides in hippocampus: evidence for a concomitant loss of synaptic proteins and interactions with an excitotoxin

Expl Neurol.

The distribution of cathepsin D activity in adult and aging human brain regions

J. Neurochem.

Cytosolic proteolysis of tau by cathepsin D in hippocampus following suppression of cathepsins B and L

J. Neurochem.

Selective suppression of cathepsin L results from elevations in lysosomal pH and is followed by proteolysis of tau protein

NeuroReport

Suppression of cathepsins B and L causes a proliferation of lysosomes and the formation of meganeurites in hippocampus

J. Neurosci.

Unilateral injury to the adult rat optic nerve causes multiple cellular responses in the contralateral site

J. Neurobiol.

Neuropathological staging of Alzheimer-related changes

Acta neuropath.

Pigmentoarchitectonic pathology of the isocortex in juvenile neuronal ceroid-lipofuscinosis: axonal enlargements in layer IIIab and cell loss in layer V

Acta neuropath.

Transplantation of fetal suprachiasmatic nuclei into middle-aged rats restores diurnal Fos expression in host

Am. J. Physiol.