Laboratory instruction in histology at the University at Buffalo: recent replacement of microscope exercises with computer applications.

Histology is a morphologic science in which the structure of the cells, tissues, and organs of the body are examined with a microscope. In the laboratory courses in histology at the School of Medicine of the University at Buffalo, histologic specimens had been used since the late 19th century to teach the principles of cell, tissue, and organ structure. Students also had to learn how to analyze or "read" slides with a microscope. Learning histology in this way, i.e., by direct examination of actual specimens, is time consuming and viewed by some as unnecessary. As a result of recent curricular reform at the School of Medicine that reduced contact time in histology, half of all laboratory exercises that would have been performed with a microscope were performed instead with interactive computer applications. By replacing some microscope exercises with more efficient computer applications, the histology course accommodated curricular change by both reducing contact time and continuing to offer valuable microscope laboratories for most of the organ systems of the body. To provide a basis for comparing traditional microscope exercises with computer-assisted instruction in histology, the nature of the laboratory experience between 1846 and 1998 is briefly reviewed. The instructional strategy behind the use of computers is presented, along with the nature of the computer applications and the means by which the computer applications were incorporated into the school's laboratory course in histology.

Neuropeptide Y-like immunoreactivity in the diencephalon of the little brown bat (Myotis lucifugus): localized variations with physiological state.

The current study used light microscopic immunocytochemistry to demonstrate and compare neuropeptide Y-like immunoreactivity (NPY-IR) in the diencephalon of the little brown bat (Myotis lucifugus) at different stages in its annual cycle of activity and hibernation. Animals were sacrificed in each of three discrete physiological states: euthermic, hypothermic, and hibernating. In general, NPY-IR was abundant in the hypothalamus and sparse in other diencephalic areas. Immunoreactivity was present in a number of pathways which project to or originate from diencephalic nuclei; these include the ansa peduncularis, medial forebrain bundle, inferior thalamic peduncle, stria terminalis, stria medullaris, mammillary peduncle, and dorsal longitudinal fasiculus. Dense fiber plexuses were present throughout the hypothalamus; however, NPY-IR was conspicuously absent from the suprachiasmatic nucleus. Immunoreactive perikarya were located in the supraoptic, dorsomedial, ventromedial, and arcuate nuclei, in the external division of the ventral lateral geniculate nucleus, and in the pineal gland. Localized changes in density and/or distribution of NPY-IR were correlated with changes in physiological state.

Attenuation of contraction-induced skeletal muscle injury by bromelain.

The proteolytic enzyme, bromelain, reportedly has therapeutic effects in the treatment of inflammation and soft tissue injuries. We tested the hypothesis that bromelain attenuates skeletal muscle injury induced by lengthening contractions. The left extensor digitorum longus (EDL) muscle of anesthetized hamsters was injured using a motorized foot pedal which repeatedly flexed/extended the foot through a range of 125 degrees. The EDL muscle was electrically stimulated for 400 ms during plantarflexion. Animals were assigned randomly to either a 0-d group (evaluated 3-h post-injury) or to untreated (UT) or bromelain-treated (T) groups, evaluated 3, 7, or 14 d post-injury. Following injury, T received 5 mg.kg-1 b.w. of bromelain, twice daily. Maximum isometric tetanic force (Po) was measured in vitro, then muscles were fixed, sectioned, and examined for evidence of fiber damage. The Po of injured muscles from T were higher than Po of injured muscles from UT at 3 (18.7 +/- 0.4 vs 16.5 +/- N.cm-2 and 14 d (20.5 +/- 0.6 vs 18.2 +/- 0.6 N.cm-2) (P less than 0.05), but not 7 d (19.5 +/-0.7 vs 17.7 +/- 0.8 N.cm-2). The Po of UT injured muscles were significantly lower than Po of contralateral control muscles at all time periods. Po of injured muscles from T were lower than Po from control muscles at 3 and 7 d (P less than 0.05), but not 14 d. The number of intact fibers of 3-d UT injured muscles was lower than the number of intact fibers in control muscles (P less than 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Neuropeptide Y innervation of the cerebellum in the little brown bat (Myotis lucifugus).

Light microscopic immunocytochemistry for neuropeptide Y (NPY) has demonstrated the presence of a diffuse system of NPY-immunoreactive fibers in the cerebellum of Myotis lucifugus. These fibers gain access to the cerebellum by way of the superior cerebellar peduncle, and terminate on Purkinje cell dendrites and on neurons in the granule cell layer. The origin of cerebellar NPY and possible colocalization with norepinephrine are discussed.

Cholecystokinin (CCK)-like immunoreactivity in the brain of the little brown bat (Myotis lucifugus).

The distribution of Cholecystokinin (CCK-8)-like immunoreactivity was mapped in the brain of the little brown bat, Myotis lucifugus, at three different and discrete levels of physiological activity: euthermic, hypothermic, and hibernating. Immunoreactive perikarya were present in the cerebral cortex, hippocampal formation, several nuclei of the olfactory and limbic systems, the ventral lateral geniculate nucleus, suprachiasmatic nucleus, medial geniculate nucleus, and caudate-putamen. Immunoreactive fibers were present in plexuses throughout the brain and in three major projection pathways: the medial forebrain bundle, the mammillary peduncle, and dorsal longitudinal fasciculus. Our data suggest two possible loci for CCK regulation of feeding behavior: a hypothalamic locus in the dorsomedial nucleus and a brainstem locus in the nucleus tractus solitarius.

Endocytosis of cationized ferritin by rat photoreceptors.

Endocytosis by rods of the rat retina was studied at the ultrastructural level with cationized ferritin. Endocytosed ferritin was found within all photoreceptor subdivisions except the outer segment. Within the cell, synaptic vesicles, lysosomes and a variety of other membrane-bound organelles including the axonal agranular reticulum contained ferritin. These findings indicate that surface membrane and materials bound to surface membrane are recycled to form synaptic vesicles, that a portion of the captured membrane and extracellular material enters a lysosomal pathway, and that endocytosed materials can be retrogradely transported to the myoid region of the photoreceptor.

VIP-like immunoreactivity in the anterior hypothalamic area of a hibernating bat varies with physiological state.

Vasoactive intestinal polypeptide (VIP)-like immunoreactivity in the anterior hypothalamic area (AHA) was studied in 15 bats which are seasonal hibernators, using the unlabeled antibody enzyme method of Sternberger. Our results showed that the density of immunoreactive fibers and terminal plexuses in the AHA was greatest in euthermic animals and decreased dramatically when body temperature and cardiac rate were depressed and animals entered hibernation.

Immunocytochemical localization of vasoactive intestinal polypeptide (VIP) in the brain of the little brown bat (Myotis lucifugus).

Vasoactive intestinal polypeptide (VIP)-like immunoreactivity has been examined in the brain of the little brown bat, Myotis lucifugus, using light microscopic immunocytochemistry and the indirect antibody enzyme method of Sternberger. Animals were sacrificed at three different and discrete levels of physiological activity: euthermic, hypothermic and hibernating. The density and distribution of immunoreactive neurons and fibres was compared in the three animal groups with the aid of a computerized image analysis system. Our results were compared with those of previous studies in laboratory species such as the rat and cat. Our study has demonstrated marked changes in the density of VIP-immunoreactive fibres and plexuses in the anterior hypothalamic area which correspond to the physiological state of the animal. In addition we have demonstrated the presence of VIP immunoreactive perikarya in a number of previously unreported locations. These include the paraventricular and periventricular hypothalamic nuclei, the linear raphe nucleus, nucleus interfascicularis, and in neurons embedded in the fibres of the dorsal tegmental decussation.

Distribution of somatostatinlike immunoreactivity in the brain of the little brown bat (Myotis lucifugus).

The distribution of somatostatinlike immunoreactive (SLI) perikarya, axons, and terminals was mapped in subcortical areas of the brain of the little brown bat, Myotis lucifugus, using light microscopic immunocytochemistry. A preponderance of immunoreactivity was localized in reticular, limbic, and hypothalamic areas including: 1) in the forebrain: the bed nucleus of the stria terminalis; lateral preoptic, dorsal, anterior, lateral and posterior hypothalamic areas; amygdaloid, periventricular, arcuate, supraoptic, suprachiasmatic, ventromedial, dorsomedial, paraventricular, lateral and medial mammillary, and lateral septal nuclei; the nucleus of the diagonal band of Broca and nucleus accumbens septi; 2) in the midbrain: the periaqueductal gray, interpeduncular, dorsal and ventral tegmental, pretectal, and Edinger-Westphal nuclei; and 3) in the hindbrain: the superior central and parabrachial nuclei, nucleus incertus, locus coeruleus, and nucleus reticularis gigantocellularis. Other areas containing SLI included the striatum (caudate nucleus and putamen), zona incerta, infundibulum, supramammillary and premammillary nuclei, medial and dorsal lateral geniculate nuclei, entopeduncular nucleus, lateral habenular nucleus, central medial thalamic nucleus, central tegmental field, linear and dorsal raphe nuclei, nucleus of Darkschewitsch, superior and inferior colliculi, nucleus ruber, substantia nigra, mesencephalic nucleus of V, inferior olivary nucleus, inferior central nucleus, nucleus prepositus, and deep cerebellar nuclei. While these results were similar in some respects to those previously reported in rodents, they also provided interesting contrasts.

Retinofugal projections of the big brown bat, Eptesicus fuscus and the neotropical fruit bat, Artibeus jamaicensis.

Retinal connections were studied in Eptesicus fuscus and Artibeus jamaicensis using anterograde axonal degeneration and autoradiographic techniques following unilateral enucleations and uniocular injections of radioactive amino acids. Although each retina projected bilaterally to the brainstem, the number of silver grains in the emulsion of autoradiographs indicated that nearly all fibers in the optic nerve entered the contralateral optic tract. Ipsilaterally, a major portion of the projection ended in the suprachiasmatic nucleus; caudal to the suprachiasmatic nucleus, the amount of label was so small that individual silver grains were counted to determine the location and quantity of label in other ipsilateral nuclei. In both species the retinal projection terminated bilaterally in the suprachiasmatic, dorsal lateral geniculate, ventral lateral geniculate, and pretectal olivary nuclei and contralaterally in the posterior pretectal nucleus, superficial gray layers of the superior colliculus, and nuclei of the accessory optic system. In Eptesicus the projection to the nucleus of the optic tract ended contralaterally, and in Artibeus it ended in this nucleus bilaterally. The results of this study revealed a basic theme in the optic projection of the two ecologically different microchiropterans. The results differed, however, in that the projection was larger and visually related nuclei were better developed in Artibeus. Such variations are presumed to relate to eye size and the relative use of vision by the two chiropterans.

Ultrastructure of remnant photoreceptors in advanced hereditary retinal degeneration.

The outer layers of the retinas of pigmented rats affected with hereditary retinal degeneration (rdy) were studied at an advanced stage in the degenerative process, ie, in 200 day old animals. At this age, most of the photoreceptors that survived the degenerative process were cones. The chromatin pattern of their nuclei clearly differentiated them from rods, displaced pigment cells and/or presumptive macrophages that also were found in the outer nuclear layer. None of the cones encountered had outer segments although structures resembling outer segment discs were found with a single cilium. Cones that had inner segments were found in regions of the retina that contained large accumulations of cellular debris. Cones that had lost both inner or outer segments, on the other hand, were found in regions that contained less debris. In such cells, the perikaryon of the cone was rich in mitochondria and other organelles; and the cilium arose directly from the cell body. The morphology of the cones and the fact that they were found in regions of the retina that contained different amounts of cellular debris suggested that cones with inner segments were in an early stage of degeneration while those that had lost inner segments were in a later stage of degeneration. All the cones encountered contained a variety of organelles including: free ribosomes, rough endoplasmic reticulum, and the Golgi apparatus. The cones that survived retinal degeneration therefore appeared to retain the cellular organelles needed for the production of photosensitive pigments. As a consequence, they may be capable of photoreceptor functions.

Quantitative measures of mature neuronal morphology after in utero ethanol exposure.

Dendritic networks of neocortical pyramidal cells and cerebellar Purkinje cells of mature adult rats which were exposed to ethanol in utero were analyzed for quantitative deviations from the normal branching patterns present in neurons of unexposed rats. No significant quantitative differences were found between cells of mature ethanol-exposed and control animals. The absence of quantitative deficits in the branching dendrites of these neuronal populations (which do exhibit ethanol-induced maturational retardation when examined at birth) suggests that ethanol treatment confined to the short gestation period in these animals may be of insufficient duration to produce irreversible maturational deficits in the growth pattern of dendrites.

Uncrossed retinal projections to the accessory optic nuclei in rabbits and cats.

Retinal projections to the accessory optic nuclei of rabbits and cats were demonstrated with standard autoradiographic techniques following intraocular injections of [35S]methionine and [3H]proline. In the pigmented rabbit, albino rabbit, normally pigmented domestic cat and Siamese cat the medial, lateral and dorsal terminal nuclei (MTN, LTN, and DTN, respectively) of the accessory optic system were densely labelled on the side contralateral to the injected eye. An ipsilateral projection, while clearly present in all but the Siamese cat, varied in the number of nuclei involved. In the albino rabbit, the ipsilateral projection ended in the MTN, while in the pigmented rabbit, it ended in the MTN, LTN and DTN, and in the normally pigmented domestic cat it ended in the MTN and LTN. These results indicate that the accessory optic system in rabbits and cats is more extensive than previously reported and that differences exist in the accessory optic system which may be related to genetic differences in normally pigmented and hypopigmented animals.

Organization of the retinofugal fibers to the dorsal lateral geniculate nucleus of Pteropus giganteus.

The projection of the retina to the dorsal lateral geniculate nucleus was studied following unilateral enucleations and intraocular injections of tritiated proline in the fruit bat, Pteropus giganteus. Three cell laminae (lamina 1, 2, 3) were identified in Nissl preparations. Input from the two eyes to the laminae was partially overlapping and segregated. At levels where th input was clearly segregated the input from the contralateral eye ended in the outer portions of the three laminae while the input to the laminae from the ipsilateral eye ended along the medial edge of the optic tract (lamina S) and the inner portions of the three laminae. These results indicate that the organization of the projection is similar to that described in other mammals, i.e., the terminals of input fibers from the retina form concealed terminal laminae and they are to some degree segregated to specific terminal zones within laminae.

Visual and nonvisual units recorded from the optic tectum of Gallus domesticus.

Units recorded from the optic tectum of Gallus domesticus were tested for responses to visual, auditory and somesthetic and stimuli. Units recorded from the superficial tectal layers (stratum opticum, stratum griseum et fibrosum superficiale) responded only to visual stimuli. On the basis of the unit's responsiveness to the directionality of a moving visual stimulus, three unit types referred to as pandirectional, unidirectional, and multidirectional units were identified. In the deeper layers (stratum griseum centrale, stratum album centrale) unimodal and bimodal units were encountered. These units responded to visual, auditory, and/or somesthetic stimuli. These results indicate that the tectal cortex of Gallus domesticus is subdivided into two functional divisions: the superficial layers which correlate visual information and the deep layers which correlate visual and nonvisual information.