Yasuhiro Torigoe

Retinal pathology in Alzheimer's disease. I. Ganglion cell loss in foveal/parafoveal retina

Morphometric analysis of the numbers of neurons in the ganglion cell layer (GCL) of the central retina (fovea/foveola/parafoveal retina) in eyes from 9 Alzheimers disease (AD) and 11 age-matched control cases revealed an overall decrease of 25% in total numbers of neurons in AD as compared with control eyes. Detailed analyses of GCL neurons at various eccentricities from the foveola showed that the greatest decrease in neuronal density (43% decrease) occurred in the central 0-0.5 mm (foveal region), while at 0.5-1 mm and at 1-1.5 mm eccentricities, neuronal loss amounted to 24 and 26%, respectively. The temporal region of the central retina appeared most severely affected, with up to 52% decrease in neuronal density near the foveola (central 0-0.5 mm eccentricity). There was close agreement between fellow eyes analyzed separately for three AD and three control cases. Analysis of neuronal sizes showed that all sizes of neurons were similarly affected in AD. In the GCL of control retinas, neurons decreased with age (coefficient of correlation = -0.67), while in AD retinas no such relationship was evident. Since in the central 0-2 mm region of the retina 97% of neurons in the GCL are ganglion cells (while the remaining 3% consist of displaced amacrine cells), these results demonstrate extensive ganglion cell loss in the central retina in AD.

Retinal pathology in Alzheimer's disease. II. Regional neuron loss and glial changes in GCL.

Detailed analyses of neuronal and astrocyte cell numbers in the ganglion cell layer (GCL) of whole-mounted peripheral retinas from 16 Alzheimers disease (AD) and 11 control eyes (11 and 9 cases, respectively) demonstrate extensive neuronal loss throughout the entire retina in AD as compared to control eyes. The observed neuronal loss is most pronounced in the superior and inferior quadrants, ranging between 40 and 49% throughout the midperipheral regions, and reaching 50-59% in the far peripheral inferior retina, while the overall neuronal loss throughout the entire retina amounts to 36.4% (p < 0.004). Although the 16% increase in astrocyte numbers is not significant, the ratio of astrocytes to neurons is significantly higher (82%; p < 0.0008) in AD as compared to normal retina (0.238 +/- 0.070 vs. 0.131 +/- 0.042). These results are strengthened by the close agreement (within +/- 15% of respective means) found between fellow eyes. Analysis of glial fibrillary acidic protein immunoreactivity (GFAP-ir) in sections of retinas from an additional 12 AD and 19 control cases show increased GFAP-ir with more extensive labeling of astrocytes in the GCL as well as increased labeling of Müller cell end-feet and radial processes in AD as compared to control retinas. The extensive loss of neurons documented in these retinas, accompanied by an increased astrocyte/neuron ratio, provides further support for the substantial involvement of the retina in AD.

Sensory neurons of the rat sciatic nerve

Experiments have been undertaken in this laboratory over recent years to accurately determine the numbers and sizes of somatic neurons which contribute to the normal sciatic nerve, at mid-thigh levels, of the adult, albino rat. This article is concerned with the dorsal root ganglion (DRG) neuron population of the sciatic nerve whose cell bodies were identified through retrograde labeling of cut branches of the sciatic with horseradish peroxidase (HRP) and/or its wheat germ conjugate (WGA-HRP). It is essential to understand the neuronal composition of the normal rat sciatic nerve if the consequences of aging, nerve injury, and surgical repair to improve functional regeneration are to be properly evaluated. Neuron counts were determined from camera-lucida paper drawings of all labeled profiles in DRGs L3-L6 at 100 x magnification. The profiles, obtained by labeling individual branches of the sciatic nerve (sural, lateral sural, tibial, peroneal, medial, and lateral gastrocnemius/soleus nerves) were traced from 40-microns-thick, serial, frozen sections. The sizes of the perikarya, areas and diameters, were determined by tracing the perimeters of the drawn profiles on a digitizing tablet. The tablets output was inputted directly into a specially designed computer spreadsheet which contained a mathematical table for correcting the split-cell error inherent to the sectioning process. Afferents from any given branch of the sciatic normally occupied two to three adjacent ganglia. Sciatic DRG neurons were normally located in lumbar ganglia L3-L6. Nearly 98-99% of all sciatic DRG perikarya resided in the L4 and L5 DRGs. The L6 DRG, traditionally regarded as an important contributor to the rat sciatic, contained merely 0.4% of its afferent neurons while the L3 ganglion, frequently overlooked as a contributor, contained 1.2% of the mid-thigh sciatic afferents. The mean size of rat DRG neurons was about 29 microns (550-600 microns2). The corrected counts revealed that the normal sciatic nerve (at mid-thigh levels), in rats between 2 and 12 months of age, contained a mean, total DRG neuron population of about 10,500 neurons. This is probably an underestimate by 3-5% of the true number due to occasional unreliable labeling of some of the small DRG neurons. It is estimated that the normal, mean number of sciatic DRG neurons of young to middle-aged rats lies somewhere between 10,500 and 11,000 +/- 2000. The data suggest that nearly 20% of all DRG neurons in the sciatic nerve supply muscle afferents. The vast majority of the remaining neurons are involved with innervation of the skin.(ABSTRACT TRUNCATED AT 400 WORDS)

Afferent projections to the cerebellar flocculus in the pigmented rat demonstrated by retrograde transport of horseradish peroxidase.

SummaryThe horseradish peroxidase (HRP) retrograde transport method was used to identify brainstem afferents to the cerebellar flocculus in the pigmented rat. Injections of the enzyme were made through recording microelectrodes, making it possible to localize the injection site by physiological criteria. Clearly, the largest number of afferents arise from the bilateral vestibular and perihypoglossal nuclei and from the contralateral dorsal cap (of Kooy) of the inferior olive. Additionally, a substantial number arise bilaterally from: (1) the nucleus reticularis tegmenti pontis (NRTP); (2) several of the cranial motor nuclei including the abducens, retrofacial and facial nuclei and the nucleus ambiguus; (3) the rostral part of the lateral reticular nucleus (subtrigeminal nucleus); (4) the raphe pontis and raphe magnus and (5) neurons intercalated among the medial longitudinal fasciculus (MLF) just rostral to the hypoglossal nucleus and another group rostral to the abducens nucleus.The basilar pontine nuclei contained a large number of lightly labeled neurons in all flocculus injections which were discretely located within the dorsolateral, lateral and medial divisions. These areas were labeled bilaterally but with a slight contralateral preponderance. Injection into the flocculus, but involving the adjacent ventral paraflocculus, produced a heavier labeling of pontine neurons with a slightly different distribution. Therefore, we tentatively conclude that the flocculus receives input from these pontine visual centers (dorsolateral, lateral and medial nuclei), perhaps through collateral projections from neurons projecting to the paraflocculus.The present study demonstrates strong similarities between the rat and other species studied (e.g., rabbit, cat, monkey) in terms of the brainstem nuclei projecting to the flocculus. Most noticeable in quantitative terms are the pathways known to mediate vestibular (vestibular and perihypoglossal nuclei) and visual (optokinetic) information (e.g., NRTP). Additionally, we can provide morphological evidence that the midline and paramedian pontine tegmentum, identified in the cat and monkey as containing saccade-related neurons, send large numbers of projections to the rat flocculus. Given these similarities, the rat may be a suitable animal model in which to study the pathways underlying visual-vestibular interaction and saccadic mechanisms in the flocculus.

Retinal degeneration in the macula of patients with Alzheimer's disease.

Recent reports (Hinton et al. 1986; Blanks et al. 1989) document the involvement of the retina in the constellation of neurodegenerative changes present in Alzheimers disease (AD). These studies demonstrate the degeneration of large numbers of optic nerve axons and loss of retinal ganglion cells (RGCs) in patients with AD, but the quantitative changes in the retina of patients with AD compared with age-matched controls have not been examined. An important question is whether the lesion affects the macula, the area of highest visual acuity and the region of the greatest density of cone photoreceptor cells and RGCs. Additionally, it is unknown if patients with AD have a uniform thinning of cells in the ganglion cell layer (GCL) or if there is a differential loss of the medium- to large-sized cells, as suggested earlier (Bassi et al. 1987) and documented histopathologically in some areas of the central nervous system of patients with AD (Kemper 1984). If patients with AD were to show a differential loss of large versus small RGCs with characteristic differences in density, distribution, central projections, and physiologic properties (see review by Rowe and Stone 1977), then a loss of the visual functions normally ascribed to these classes of mammalian RGCs might be expected. This quantitative study of the retinal lesions in the macula of patients with AD provides important data on the progression of the disease and may eventually be the basis for diagnostic procedures for assessing the severity of AD.

Orientation of the semicircular canals in rat

The orientation of the rat semicircular canals was determined using one of two techniques. Null point analysis was used to define physiologically the planar equations of the anterior (n = 15) and posterior canals (n = 15); equations for the horizontal canal (n = 19) were determined using an anatomical dissection technique. Canal orientation was defined with respect to stereotaxic coordinate system and, for comparison, relative to head position during freeze (startle) behavior. Results show that ipsilateral canal planes are orthogonal within 4-8 degrees, and pairs of right-left synergistic pairs are essentially co-planar. The horizontal canals are inclined upwards 35 degrees with respect to the horizontal plane, but a head position of 43 degrees nose-down was determined to produce near optimal horizontal canal and minimal vertical canal activation with horizontal rotation. Finally, a loud or unexpected auditory stimulus initiates a freeze (startle) response in rat characterized by an transient followed by a sustained head position lasting several seconds. Transients are complete within 300-400 ms. Thereafter, the head becomes momentarily stabilized in the startle position which averaged 14 +/- 8 degrees (nose-down with respect to horizontal stereotaxic zero) across the population (n = 14). The response habituated only slightly, but the final position was sufficiently variable so as to limit the usefulness of the freeze (startle) position as a reference of semicircular canal position in the rat.

Projections of the nucleus of the optic tract to the nucleus reticularis tegmenti pontis and prepositus hypoglossi nucleus in the pigmented rat as demonstrated by anterograde and retrograde transport methods

The visual pathways from the nucleus of the optic tract (NOT) to the nucleus reticularis tegmenti pontis (NRTP) and prepositus hypoglossi nucleus (ph) were studied following injections of tritiated leucine into the NOT of pigmented rats. The cell bodies of origin of the pretectal-NRTP, NRTP-ph, and pretectal-ph projections were determined using retrograde horseradish peroxidase (HRP) technique. The pretectum projects strongly to the rostral two-thirds of the central and pericentral subdivisions of the NRTP and sends a remarkably smaller projection to the ph. Both are entirely ipsilateral. The fibers destined for the ph travel with the NOT-NRTP bundle, pass through the NRTP, traverse the medial longitudinal fasciculus, and are distributed to the rostral one-half of the ph. The retrograde HRP studies confirm these pathways. The pretectal projections to the NRTP arise from neurons in the rostromedial NOT; those to the ph are located primarily in the rostral NOT although small numbers are found within the anterior, posterior, and olivary pretectal nuclei. Of major importance is the fact that the ph injections retrogradely label neurons within the NRTP and the adjacent paramedian pontine reticular formation. This NRTP-ph projection is entirely bilateral and arises from parts of both subdivisions of the nucleus targeted by NOT afferents. Both the direct NOT-ph and indirect NOT-NRTP-ph connections provide the anatomical basis for the relay of visual (optokinetic) information to the perihypoglossal complex and, presumably, by virtue of reciprocal ph-vestibular nuclear connections, to the vestibular nuclei itself. Such pathways confirm previous physiological studies in rat and, in particular, clarify the contrasting effects of electrolytic lesions of NRTP in rat which completely abolishes optokinetic nystagmus (OKN) (Cazin et al., 1980a) vs kainic acid lesions which produce only minor effects on OKN slow velocity (Hess et al., 1988). Given these differential effects, one concludes that the critical pathway for OKN passes in relation to, but is not significantly relayed by, the neurons of the NRTP or adjacent pontine tegmentum. The present studies suggest that one such fiber system is the NOT-ph bundle. How this relatively small projection compares to other possible fiber of passage systems remains to be determined electrophysiologically.

Sympathetic preganglionic efferent and afferent neurons mediated by the greater splanchnic nerve in rabbit

Motion sickness, a multisymptom disorder characterized by abnormal gastrointestinal motility and emesis, can be induced by vestibular effects on the sympathetic portion of the autonomic nervous system. However, the vestibular-autonomic pathways are unknown. As a first step in the analysis, we identified the locus of preganglionic sympathetic neurons (PSNs) and dorsal root afferent ganglionic neurons (DRGs) which supply sympathetic innervation to major portions of the gastrointestinal tract in the rabbit. Retrograde labeling of neurons was obtained by application of horseradish peroxidase (HRP) to the cut end of the greater splanchnic nerve. Labeled PSNs were found, ipsilaterally, within the T1 to T11 spinal cord segments, with the highest density of neurons in T6. Most PSNs were located within the intermediolateral column (IML), but a significant portion also occurred within the lateral funiculus (LF), the intercalated region (IC) and the central autonomic area (CA). The proportion of labeling between the four regions depended on the spinal cord segment. In the midthoracic levels, the distribution of labeled neurons was denser in the IML and LF, and in the caudal thoracic segments, the majority were localized in the IC and CA. Labeled cells in these four areas varied morphologically from large fusiform neurons in the IC to small fusiform neurons in the LF, small stellate neurons in the CA, and medium-size stellate neurons in the IML. The DRGs were labeled in thoracic segments T1 to T12, with the majority between T5 and T11. These labeled DRG somata of the greater splanchnic nerve were smaller in comparison with unlabeled ones.

Neurons of the medial terminal accessory optic nucleus of the rat are poorly collateralized

The vast majority of neurons of the rat medial terminal nucleus (MTN) project to the nucleus of the optic tract (NOT), but the MTN also projects to a lesser degree upon a number of other brainstem nuclei controlling optokinetic nystagmus. Because of the diversity of targets of the MTN, it is possible that individual neurons have branched axons that project to two or more brainstem nuclei. The possibility that axons of MTN-NOT neurons collateralize to innervate other MTN targets is examined in the rat with the fluorescent, double-labeling, retrograde tracer technique. Fluoro-Gold was injected into the NOT while Fast Blue was simultaneously injected into each of five other known targets of the MTN: the supraoculomotor-periaqueductal gray; the dorsal cap of the inferior olive; the visual tegmental relay zone; the dorsolateral nucleus of the basal pons; and the superior/lateral vestibular nuclei. Brainstem sections were processed for fluorescence microscopy and the MTN was examined for single- and double-labeled neurons. Results show that virtually all neurons of the MTN (greater than 97.5%), together with neurons in the visual tegmental relay zone immediately surrounding the MTNd, are single-labeled in all paired injections involving the NOT and the other target nuclei. It was found that about 69% of MTN neurons project exclusively to the NOT, 5-10% project to each one of the other nuclei, and 3% of MTN neurons project to more than one target. Based upon cell counts from the fluorescent material, and previous analysis of Nissl-stained serial sections, the findings show that virtually all MTN neurons are projection neurons. It was concluded that the MTN is comprised of independent projection systems, possibly involved in different aspects of generating optokinetic nystagmus.

Opioid receptors in the accessory optic system of the rat: effects of monocular enucleation.

The presence and concentrations of each of the three subtypes of opioid receptors (mu, kappa, and delta) has been studied in the accessory optic nuclei (dorsal, lateral, and medial terminal nuclei and the interstitial nucleus of the superior fasciculus, posterior fibers: DTN, LTN, MTN, and inSFp) in normal young rats with radioligands directed towards each opioid receptor subtype. The changes in mu opioid receptors have also been investigated in monocularly enucleated rats in which one eye was removed and the rats sacrificed at postoperative day (PO) 2, 3, 5, 7, 14, and 30. As the MTN is the only accessory optic nucleus of the rat large enough for semiquantitative evaluation, the mu receptor population of the MTN has been subjected to optical microdensitometric analysis. All four of the accessory optic nuclei (AOS nuclei) are found to contain exceedingly high levels of mu opioid receptor binding with the selective radioligand [3H]-[D-Ala,MePhe4, Gly-ol5] (DAGO), low levels of kappa opioid receptor binding using the radioligand [3H]-[ethylketocyclazocine] (EKC) together with the competing agents [D-Pro4]-morphiceptin and [D-Ser2,Thr6]-Leu-enkephalin, and an absence of delta opioid receptor binding with the radioligand [3H]-[D-Ala2,D-Leu5]-enkephalin (DADLE) combined with the competing agent [D-Pro4]-morphiceptin. Monocular enucleation, as studied on the mu opioid receptor population with this experimental approach, results in virtually a complete loss of mu opioid receptors throughout all four of the contralaterally located AOS nuclei, including both dorsal and ventral subdivisions of the medial terminal nucleus (MTNd,v). Kappa and delta receptors are very few (kappa receptors) or are lacking (delta receptors) in the AOS nuclei, and for this reason, the effects of monocular enucleation on these two opioid receptor subtypes have not been investigated. Monocular enucleation also produces a significant lowering in mu receptor binding in other primary optic nuclei (the lateral geniculate nuclei, nucleus of the optic tract, and superficial layers of the superior colliculus) and in the pars principalis of the medial geniculate nucleus (description of changes in mu receptors in non-accessory optic primary optic nuclei will be considered elsewhere). Microdensitometric study of the MTNd,v shows that the decreased binding of mu receptors in this nucleus is barely detectable (about 6%) at PO2 and rises to 6-15% at PO3. At PO5 receptor loss reaches approximately 62%, whereas at PO7 it is about 81% complete. At PO14 and PO30, the mu receptor loss is nearly complete at around 93%.(ABSTRACT TRUNCATED AT 400 WORDS)