Microcebus murinus: a useful primate model for human cerebral aging and Alzheimer's disease?

Age-associated dementia, in particular Alzheimer's disease (AD), will be a major concern of the 21st century. Research into normal brain aging and AD will therefore become increasingly important. As for other areas of medicine, the availability of good animal models will be a limiting factor for progress. Given the complexity of the human brain, the identification of appropriate primate models will be essential to further knowledge of the disease. In this review, we describe the features of brain aging and age-associated neurodegeneration in a small lemurian primate, the Microcebus murinus, also referred to as the mouse lemur. The mouse lemur has a relatively short life expectancy, and animals over 5 years of age are considered to be elderly. Among elderly mouse lemurs, the majority show normal brain aging, whereas approximately 20% develop neurodegeneration. This Microcebus age-associated neurodegeneration is characterized by a massive brain atrophy, abundant amyloid plaques, a cytoskeletal Tau pathology and a loss of cholinergic neurons. While elderly mouse lemurs with normal brain aging maintain memory function and social interaction, animals with age-associated neurodegeneration lose their cognitive and social capacities and demonstrate certain similarities with age-associated human AD. We conclude that M. murinus is an interesting primate model for the study of normal brain aging and the biochemical dysfunctions occurring in age-associated neurodegeneration. Mouse lemurs might also become an increasingly important model for the development of novel treatments in this domain.

Fibrous Histiocytoma-like Spindle-Cell Proliferation in the Nipple After Body-Piercing.

We report the case of a 19-year-old pregnant woman who presented with a nipple tumor. The lesion consisted in a spindle-cell proliferation with histologic features similar to those of fibrous histiocytoma, with a highly vascularized stroma. Although it showed low mitotic activity, scattered marked atypical cells with prominent nucleoli were identified, thus raising concern about the benign nature of the tumor. Immunohistochemical evaluation revealed that the spindle cells were diffusely positive for vimentin, focally positive for CD68, and negative for all the other tested antibodies. The patient had a total excision of the lesion and she is free of disease after 30 months. To our knowledge this is the first reported case of a lesion of this type in the nipple after body-piercing.

Senile plaques and neurofibrillary changes in the brain of an aged lemurian primate, Microcebus murinus.

In some aged Microcebus brains (8- to 11-year-old animals) dramatic atrophy is found, particularly of the cortex, the hippocampus, the basal ganglia, the brainstem and the cerebellum, associated with a conspicuous increase in the size of the cerebral ventricles. These morphological changes are accompanied by certain histological profiles indicative of pathology. In the cortex, these histological changes consist of 1) a large number of senile plaques composed of degenerated neurites sometimes surrounding an amyloid plaque, 2) amyloid deposits in the vascular walls and 3) dense bundles of argyrophilic filaments in numerous pyramidal neurons. All these lesions resemble changes associated with Alzheimer's disease in man. The degenerative changes observed in the Microcebus brain are accompanied by behavioral changes. At the moment these preliminary studies, carried out on the smallest of all primates, do not prove that the degeneration is of the Alzheimer type, but do indicate that Microcebus murinus may well be a good model for the study of cerebral aging, providing a comparison with cerebral ageing in humans. The size, life span and cost of the animal provide further advantages when compared with other nonhuman primates.

Identification of amyloid beta protein in the brain of the small, short-lived lemurian primate Microcebus murinus.

The deposition of amyloid beta (A beta) protein in the brain has been demonstrated immunocytochemically in the small Lemurian primate Microcebus murinus. Both meningocerebral vascular deposits and cortical parenchymal deposits occur. All eight aged (> 8 years old) Microcebus examined showed vascular amyloid deposits, whereas only four exhibited parenchymal plaques. The vascular amyloid infiltrated the tunica media of the leptomeningeal and cortical arteries and arterioles and was also found in capillaries. A beta was observed to be deposited in three general forms in the cortical neuropil: round or elliptical plaques that were thioflavin-negative but sometimes showed a central concentration of A beta immunoreactivity; round plaques with a densely immunoreactive core that was thioflavin-positive; extensive ribbon-like infiltrations enclosing multiple cortical blood vessels. These observations, taken together with previous descriptions of age-related neurodegenerative changes in Microcebus, indicate that this species undergoes a beta-amyloid-associated neuropathology highly similar to that seen in Alzheimer's disease. We conclude that this lemurian primate of small size and relatively short life expectancy, provides a compelling animal model of some principal features of Alzheimer's disease.

Choline acetyltransferase and somatostatin levels in aged Microcebus murinus brain.

beta-Amyloid protein (beta-AP) deposits, analoguous to those found in Alzheimer's disease (AD) are observed in the brain of aging Microcebus murinus. Because choline acetyltransferase (ChAT) activity and somatostatin (SRIH) content are consistently decreased in AD, we tested whether such changes could be observed in middle aged to aged Microcebus cerebral cortex and whether they were accompanied by beta-AP deposits. A positive correlation was observed between age and ChAT activity. By HPLC, SRIH immunoreactivity eluted as four peaks, two of which being identical with SRIH-28 and SRIH-14 while the other two likely represented precursor forms. Cortical SRIH content was not significantly affected by age. ChAT activity and SRIH content were not significantly correlated. Amyloid angiopathy was observed in every brain examined and the presence of cortical lesions analoguous to senile plaques observed in the oldest case only which did not demonstrate important alterations in ChAT and somatostatin levels.

Localization and characterization of neuropeptide Y in the brain of Microcebus murinus (Primate, Lemurian).

The distribution of neuropeptide Y (NPY) in the brain of the lemur Microcebus murinus was determined by immunocytochemistry with the aid of a highly specific antiserum against synthetic porcine NPY. When compared with previous immunohistochemical data obtained in primates and other mammalian species, the localization of NPY-immunoreactive (IR) structures in the Microcebus murinus brain revealed particular features. (1) Numerous NPY-IR perikarya and a dense network of IR nerve terminals were found in the supraoptic and suprachiasmatic nuclei, respectively. The occurrence of NPY-IR perikarya in the supraoptic nucleus, also reported in the squirrel monkey, seems to be specific to primates. In the squirrel monkey, the suprachiasmatic nucleus exhibits only a moderate innervation, whereas in humans it appears totally devoid of NPY-IR fibers. (2) IR perikarya and axon processes were observed in many upper brainstem areas, in particular in the interpeduncular, raphe pontine, dorsal tegmental, parabrachial, and dorsal raphe nuclei, in the locus coeruleus, the nucleus of the solitary tract, and the reticular formation; in this latter area, the occurrence of two categories of NPY-IR neurons was demonstrated on the basis of their morphology and localization, suggesting that they may play distinct roles. (3) NPY-IR nerve processes could be traced over a long distance. (4) For the first time, numerous NPY-IR terminals were observed close to the lumen of the various cerebral ventricles. The immunoreactive NPY-like peptide was characterized by combining high performance liquid chromatography (HPLC) analysis and radioimmunoassay quantification. The dilution curves obtained with synthetic porcine NPY and serial dilutions of occipital cortex, paraventricular and supraoptic hypothalamus, posterior hypothalamus, medulla oblongata, or preoptic area extracts were parallel. The highest amounts of NPY were measured in the hypothalamus and telencephalon. HPLC analysis resolved a single peak of NPY-like immunoreactivity that exhibited the same retention time as synthetic porcine NPY. The distribution of NPY in the lemurian brain is discussed with respect to phylogeny and putative functions.

Spontaneous activity of rat dorsal horn cells in spinal segments of sciatic projection following transection of sciatic nerve or of corresponding dorsal roots.

Natural forms of stimulation were used to compare the spontaneous and evoked activity of dorsal horn neurons in three groups of rats: controls with no surgical lesion, rats with transection of the sciatic nerve and rats with transection of the dorsal roots at the same segmental level. In control rats, cells encountered in the dorsal horn were classified according to their peripheral field as tactile specific, convergent tactile and nociceptive, nociceptive, or movement driven. In 20 control animals, only 20% of the 140 cells with a peripheral field were spontaneously active. After sciatic nerve transection made on the side of recording a few days previously (18 rats), all of the 141 cells studied showed spontaneous activity, only 69 of them having a peripheral field. After dorsal root transections a few days previously (nine rats), 25 spontaneously active cells were found in the dorsal horn ipsilateral to the section, none with a peripheral field. Spontaneous activities of cells without a peripheral field were separated into three types as a function of bursting pattern, which were similar following both types of transection. The spontaneous activity shown by dorsal horn cells without peripheral fields following dorsal root transection precludes attribution of spontaneous spiking in such cells to abnormal input from the periphery, and shows that abnormal activity can develop in deafferented dorsal horn cells themselves. A possible role played by this spontaneous activity in deafferentation pain is considered.

Topographical distribution of CRF immunoreactivity in the pigeon brain.

The distribution of corticotropin-releasing factor (CRF) immunoreactivity was demonstrated by immunocytochemistry in intact and colchicine-treated pigeons. Colchicine injections were administered at different times related to the circadian activity of the CRF-adrenocorticotropin (ACTH)-corticosterone axis. Three CRF antisera were used, two directed against synthetic rat CRF and one directed against synthetic ovine CRF. No fundamental differences appeared in the pigeon brain with respect to the specific CRF antiserum used. The most effective colchicine injection times corresponded to hypersecretion in the corticotropic axis. CRF-immunopositive neurons were scattered throughout the pigeon brain. In addition to the paraventricular hypothalamic system, which is involved in adenohypophysial ACTH regulation, several other hypothalamic and extrahypothalamic areas showed CRF neurons. The distribution suggests that CRF may also act as a modulator and a neurotransmitter. Two hypothalamic paraventricular nucleus-median eminence CRF pathways are described here. Moreover, CRF-immunopositive reactions were observed in specific areas of cerebral ventricle walls, suggesting that CRF may be released into the cerebral fluid.

A stereotaxic atlas of the grey lesser mouse lemur brain (Microcebus murinus).

In response to the growing interest in the prosimian Microcebus murinus for studies on cerebral aging, the stereotaxic atlas of its brain was carried out in view of further anatomical, biochemical, electrophysiological, and behavioral investigations as well as for therapeutic experiments. This primate, which could be a valuable model for neuroscientific studies in various domains, presents numerous physiological advantages (e.g., size, cost, and ability to breed) compared to rodents, which can be used as nonprimate models, and simians. The atlas, valid for adult microcebes of every age and both sexes, consists of 54 frontal plates and 28 sagittal plates. For the establishment of stereotaxic coordinates and for drawings and photographs, 10 adult specimens of Microcebus murinus were used. The brains were frozen, cut into sections of 50 microm thickness, every fourth section being stained with Nissl. First, sections were projected and the outlines of the different structures, nuclei, and fibers were drawn. Then, the accuracy of the analysis was improved by detailed observation directly by microscope and also by computer analysis. Finally, the photographs of the sections were scanned and processed using the software Photoshop and Illustrator. For testing coordinates, several verifications were made. Experiments on lesions and injections of different substances were carried out in specific regions of the brain and brains implanted with needles were fixed in formol and embedded in paraffin wax.

Tissue localization of overproduced esterases in the mosquito Culex pipiens (Diptera: Culicidae).

We have investigated the tissue distribution of overproduced esterases A (A1 and A2) and B (B1 and B2) in strains of Culex pipiens L. by immunocytochemistry. S-LAB mosquitoes, lacking overproduced esterases, were used as reference. Tissues showing a strong specific reaction (fluorescence) were observed with anti-esterase A1 antiserum in S54 (with A1) and BOUAKE (with A2) strains, and with anti-esterase B1 antiserum in TEM-R and EDIT (with B1) and BOUAKE (with B2) strains. Overproduction of esterases A and B was tissue-specific. The most constant pattern for the two types of esterases was their overproduction in the alimentary canal and Malpighian tubes, although fluorescence varied in intensity depending on strains and developmental stages. There was no difference in the tissue distribution of esterases Al and A2. In contrast, esterases B pattern was highly variable among strains. Differences between TEM-R and EDIT were explained by the different overall overproduction and number of copies of the amplified gene (10-fold higher in TEM-R). The most striking difference in esterase B1 and B2 tissue localization concerned the nervous system where neurons were intenisely fluorescent in TEM-R and EDIT (B1), but not in BOUAKE (B2). All esterase B positive tissues in TEM-R contained large quantities of esterase B1 mRNA (in situ hybridization), indicating that at least part of the protein revealed by immunochemistry was produced in the tissues where it was observed. Our results are discussed in terms of the protection that the different esterases can confer during exposition to organophosphorous insecticides.

Astroglial interlaminar processes in the cerebral cortex of prosimians and Old World monkeys.

Previous observations have shown that astrocytes with interlaminar processes are present in the cerebral cortex of humans and New and Old World monkeys, but not in the rodent. The present report furthers the analysis of possible evolutionary aspects regarding the expression of such astroglial features. A comparison between young and adult Microcebus murinus, a prosimian, and Old World monkeys (Macaca mulatta and Papio hamadryas) is presented. Brain samples were processed for glial fibrillary acidic protein (GFAP), vimentin, MAP2 and SMI 311 immunocytochemistry, using different procedures. The cerebral cortex of adult Microcebus showed the presence of long astroglial processes, albeit reduced in number and length with respect to those observed in Old World monkeys. Macaca and Papio showed dense packing of such processes extending in most cortical regions to a depth of approximately 700 micrometers. Based on double immunolabelling for GFAP and MAP 2 antigens, the location and extent of these processes was shown to overlap with areas traversed by bundles and individual apical dendrites. Aged Old World specimens depicted an increased thickness of terminal portions of interlaminar processes, with increased morphological alterations. Comparisons made between the average thickness of the "brush" composed of interlaminar processes and the thickness of lamina I among the species analyzed disclosed an absence of relationship between them. This suggests that interlaminar processes do not represent cellular adaptations to the increase in thickness in superficial cortical laminae, but rather to some other evolutionary pressure. Since astroglial interlaminar processes are already present in a prosimian, although in a comparatively reduced manner, it is suggested that such processes underwent an early expression within the primate order, with increasing presence in more recent primate species.

[Demonstration of centrifugal fibers in the optic chiasma and optic nerves in the rat after lesions of the suprachiasmatic nucleus]. / Mise en évidence de fibres centrifuges dans le chiasma et les nerfs optiques du rat après lésion d'un noyau suprachiasmatique.

After an unilateral destruction of the suprachiasmatic nucleus and with the use of several silver impregnation techniques, degenerating centrifugal fibers were found in both optic nerves. Centrifugal fibers to the retina originate from three different regions of the nucleus and their position in the chiasma are different. In large majority the degenerating fibers were located at the periphery of the optic nerve and were more frequent on the contralateral than on the ipsilateral side to the destroyed suprachiasmatic nucleus. The possibility that our experimental procedure demonstrates the existence of fibers originating not only in the suprachiasmatic nucleus but also in other structures whose efferent fibers pass at this level, is discussed.

Immunocytochemical identification of the mesotocin- and vasotocin-producing systems in the brain of temperate and desert lizard species and their modifications by cold exposure.

Using immunofluorescent techniques, mesotocin (MT) and vasotocin (VT) neurosecretory systems were identified in the brain of different lizards: one temperate species living in southern France and three desert species coming from southern Algeria. In the four species, MT and VT were shown to be synthesized in specific neurons located in the anterior preoptic area (POA), the supraoptic (SON), paraventricular (PVN), and ventromedial (VMN) nuclei. The neurosecretory axons of the POA neurons terminated in the vicinity of the lamina terminalis; in the three desert species, several additional VT fibers extended more rostrally, going into the olfactory bulb. The axons originating in SON, PVN, and VMN ended either in the external zone of the rostral median eminence (ME) near the adenohypophyseal portal vessels, or in the neural lobe of the hypophysis. A short exposure to cold (4 degrees) in some specimens of Lacerta muralis induced a differential response in the dorsal and ventral parts of the PVN. Whereas the dorsal part remained unchanged, the ventral part instead appeared to be essentially composed of small neurons void of secretory granules. Cold exposure also led to a marked accumulation of both MT and VT in the internal zone of the ME, but of only VT in the external ME zone. On the other hand, a conspicuous amount of MT appeared in adenohypophyseal cells.

The topography of mesotocin and vasotocin systems in the brain of the domestic mallard and Japanese quail: immunocytochemical identification.

The neurosecretory systems producing mesotocin (MT) and vasotocin (VT) (the avian homologues of oxytocin and vasopressin, respectively) were characterized in the brains of the domestic mallard and Japanese quail by means of indirect immunofluorescence techniques using specific antisera. In the anterior preoptic region, including the organum vasculosum of the lamina terminalis, and at different levels of the supraoptic and paraventricular nuclei, separate mesotocin- and vasotocin-producing neurons were identified. Mesotocinergic and vasotocinergic neurons were also located in the tuberomammillary area, among the ectomammillary tract fibers. The supraoptico-neurohypophysial tract, formed by vasotocin- and mesotocin-containing axons, enters the internal zone of the median eminence and ends in the posterior lobe of the pituitary. The external zone of the rostral median eminence appears to contain vasotocin and mesotocin fibers, which terminate in close contact with the capillaries of the hypophysial portal system.

[Immunocytochemical characterization of vasotocin and mesotocin secretory systems in duck brain]. / Caractérisation immunocytochimique des systèmes neurosécréteurs à mésotocine et à vasotocine dans l'encéphale du canard.

The neural systems secreting vasotocin and mesotocin has been characterized in the Duck brain with indirect immunofluorescent techniques, using specific antisera. In the anterior preoptic region and in supraoptic and paraventricular nuclei, neurons producing vasotocine and neurons producing mesotocine have been identified separately. Only vasotocinergic neurons were localized in ectomammillary tract. Vasotocin--and mesotocin--containing axons together enter the median eminence, some of them crossed the internal zone of the median eminence before ending in the posterior lobe of the pituitary, whereas other axons of both classes entered the external layer of the rostral median eminence, in close contact with the capillaries of the hypophysial portal system.

Retinohypothalamic pathway in the duck (Anas platyrhynchos).

Retinohypothalamic connections were studied in the duck after unilateral optic nerve transection using both light and electron microscopic techniques. Degenerated endings of optic fibers were found only in a circumscribed part of the anterior hypothalamic area, i.e. the ventral region of the contralateral suprachiasmatic nucleus. Images of degenerating boutons were observed in frozen sections (method according to Johnstone-Bowsher), and their presence confirmed by electron microscopic examination. These degenerating boutons make synaptic contacts with dendrites or dendritic spines of neurons of the suprachiasmatic nucleus. In the same material, the decussation of the optic chiasma was studied with the light microscope. Uncrossed retinal fibers were found in the marginal optic tract, the basal optic root and occasionally also inthe isthmo-optic tract.

[Demonstration of direct olfactory projections in the hypothalamus, the mesencephalon and the metencephalon of Microcebus murinus (Primate, Lemurian)]. / Mise en évidence de projections olfactives directes dans l'hypothalamus, le mésencéphale et le métencéphale de Microcebus murinus (Primate, Lémurien).

The efferent projections of the olfactory bulb in Microcebus murinus were identified after transection of the olfactory peduncle and after the revelation of the degenerating fibers by different silver staining methods. Total and partial sections have allowed demonstrating the importance of the two olfactory tracts in the olfactory projection areas. Degenerated fibers and endings were evidence not only in the different telencephalic regions, as classically known, but also in various hypothalamic nuclei (lateral hypothalamus, the suprachiasmatic, posterior supraoptic and mammillary nuclei and in the median eminence) and in several mesencephalic and metencephalic nuclei (ventral tegmental area, interpeduncular and raphe nuclei, and locus coeruleus). In all these structures the degenerate fibers were seen on both sides. The olfactory projections appeared not to be limited to the telencephalic areas. Moreover, the olfactory bulbs seem to be directly connected especially with the vegetative and integrative areas localized in the hypothalamus and the brainstem and particularly with the major aminergic nuclei that play an essential role in the neurovegetative, neuroendocrine and behavioral regulations.

[Retinal afferents of hypothalamic origin in a prosimian primate: Microcebus murinus. Study using retrograde fluorescent tracers]. / Afférences rétiniennes d'origine hypothalamique chez un Primate Prosimien: Microcebus murinus. Etude à l'aide de traceurs fluorescents rétrogrades.

The central efferent connections to the retina in Microcebus were identified using the retrograde axonal transport of fluorescent dyes. After unilateral intraocular injection of the tracer, two populations of labeled neurons were observed. Fluorescent neurons were located immediately posterior to the optic chiasma, in the anterior part of the arcuate nuclei. In addition, fluorescent neurons were observed in the suprachiasmatic nuclei; the latter pathway appears to close up an anatomical retino-hypothalamo-retinal loop between the suprachiasmatic nucleus and the retina.