Small-scale environmental enrichment and exercise enhance learning and spatial memory of Carassius auratus, and increase cell proliferation in the telencephalon: an exploratory study

Carassius auratus is a teleost fish that has been largely used in behavioral studies. However, little is known about potential environmental influences on its performance of learning and memory tasks. Here, we investigated this question in C. auratus, and searched for potential correlation between exercise and visuospatial enrichment with the total number of telencephalic glia and neurons. To that end, males and females were housed for 183 days in either an enriched (EE) or impoverished environment (IE) aquarium. EE contained toys, natural plants, and a 12-hour/day water stream for voluntary exercise, whereas the IE had none of the above. A third plus-maze aquarium was used for spatial and object recognition tests. Different visual clues in 2 of its 4 arms were used to guide fish to reach the criteria to complete the task. The test consisted of 30 sessions and was concluded when each animal performed three consecutive correct choices or seven alternated, each ten trials. Learning rates revealed significant differences between EE and IE fish. The optical fractionator was used to estimate the total number of telencephalic cells that were stained with cresyl violet. On average, the total number of cells in the subjects from EE was higher than those from subjects maintained in IE (P=0.0202). We suggest that environmental enrichment significantly influenced goldfish spatial learning and memory abilities, and this may be associated with an increase in the total number of telencephalic cells.

Microglia and neurons in the hippocampus of migratory sandpipers

The semipalmated sandpiper Calidris pusilla and the spotted sandpiper Actitis macularia are long- and short-distance migrants, respectively. C. pusilla breeds in the sub-arctic and mid-arctic tundra of Canada and Alaska and winters on the north and east coasts of South America. A. macularia breeds in a broad distribution across most of North America from the treeline to the southern United States. It winters in the southern United States, and Central and South America. The autumn migration route of C. pusilla includes a non-stop flight over the Atlantic Ocean, whereas autumn route of A. macularia is largely over land. Because of this difference in their migratory paths and the visuo-spatial recognition tasks involved, we hypothesized that hippocampal volume and neuronal and glial numbers would differ between these two species. A. macularia did not differ from C. pusilla in the total number of hippocampal neurons, but the species had a larger hippocampal formation and more hippocampal microglia. It remains to be investigated whether these differences indicate interspecies differences or neural specializations associated with different strategies of orientation and navigation.

Morphometric analysis of feedforward pathways from the primary somatosensory area (S1) of rats

We used biotinylated dextran amine (BDA) to anterogradely label individual axons projecting from primary somatosensory cortex (S1) to four different cortical areas in rats. A major goal was to determine whether axon terminals in these target areas shared morphometric similarities based on the shape of individual terminal arbors and the density of two bouton types: en passant (Bp) and terminaux (Bt). Evidence from tridimensional reconstructions of isolated axon terminal fragments (n=111) did support a degree of morphological heterogeneity establishing two broad groups of axon terminals. Morphological parameters associated with the complexity of terminal arbors and the proportion of beaded Bp vs stalked Bt were found to differ significantly in these two groups following a discriminant function statistical analysis across axon fragments. Interestingly, both groups occurred in all four target areas, possibly consistent with a commonality of presynaptic processing of tactile information. These findings lay the ground for additional work aiming to investigate synaptic function at the single bouton level and see how this might be associated with emerging properties in postsynaptic targets.

Methylmercury intoxication and histochemical demonstration of NADPH-diaphorase activity in the striate cortex of adult cats

The effects of methylmercury (MeHg) on histochemical demonstration of the NADPH-diaphorase (NADPH-d) activity in the striate cortex were studied in 4 adult cats. Two animals were used as control. The contaminated animals received 50 ml milk containing 0.42 µg MeHg and 100 g fish containing 0.03 µg MeHg daily for 2 months. The level of MeHg in area 17 of intoxicated animals was 3.2 µg/g wet weight brain tissue. Two cats were perfused 24 h after the last dose (group 1) and the other animals were perfused 6 months later (group 2). After microtomy, sections were processed for NADPHd histochemistry procedures using the malic enzyme method. Dendritic branch counts were performed from camera lucida drawings for control and intoxicated animals (N = 80). Average, standard deviation and Student t-test were calculated for each data group. The concentrations of mercury (Hg) in milk, fish and brain tissue were measured by acid digestion of samples, followed by reduction of total Hg in the digested sample to metallic Hg using stannous chloride followed by atomic fluorescence analysis. Only group 2 revealed a reduction of the neuropil enzyme activity and morphometric analysis showed a reduction in dendritic field area and in the number of distal dendrite branches of the NADPHd neurons in the white matter (P<0.05). These results suggest that NADPHd neurons in the white matter are more vulnerable to the long-term effects of MeHg than NADPHd neurons in the gray matter.

Horizontal projections of area 17 in Cebus monkeys: metric features, and modular and laminar distribution

Metric features and modular and laminar distribution of intrinsic projections of area 17 were studied in Cebus apella. Anterogradely and retrogradely labeled cell appendages were obtained using both saturated pellets and iontophoretic injections of biocytin into the operculum. Laminar and modular distributions of the labeled processes were analyzed using Nissl counterstaining, and/or cytochrome oxidase and/or NADPH-diaphorase histochemistry. We distinguished three labeled cell types: pyramidal, star pyramidal and stellate cells located in supragranular cortical layers (principally in layers IIIa, IIIb alpha, IIIb beta and IIIc). Three distinct axon terminal morphologies were found i.e., Ia, Ib and II located in granular and supragranular layers. Both complete and partial segregation of group I axon terminals relative to the limits of the blobs of V 1 were found. The results are compatible with recente evidence of incomplete segreagation of visual information flow in V 1 of Old and New World primates.

NADPH-diaphorase activity in area 17 on the squirrel monkey visual cortex: neuropil pattern, cell morphology and laminar distribution

We studied the distribution of NADPH-diaphorase activity in the visual cortex of normal adult New World monkeys (Saimiri sciureus) using the malic enzyme "indirect" method. NADPH-diaphorase neuropil activity had a heterogeneous distribution. In coronal sections, it had a clear laminar pattern that was coincident with Nissl-stained layers. In tangential sections, we observed blobs in supragranular layers of V1 and stripes throughout the entire V2. We quantified and compared the tangential distribution of NADPH-diaphorase and cytochrome oxidade blobs in adjacent sections of the supragramular layers of V1. Although their spatial distributions were rather similar, the two enzymes did not always overlap. The histochemical reaction also revealed two different types of stained cells: a slightly stained subpopulation and a subgroup of deeply stained neurons resembling a Golgi impregnation. These neurons were sparsely spined non-pyramidal cells. Their dendritic arbors were very well stained but their axons were not always evident. In the gray matter, heavily stained neurons showed different dendritic arbor morphologies. However, most of the strongly reactive cells lay in the subjacent white matter, where they presented a more homogenous morphology. Our results demonstrate that the pattern of NADPH-diaphorase activity is similar to that previously described in Old World monkeys.

Morphometric analysis of intrinsic axon terminals of Cebus monkey area 17

A morphological study of intrinsic projections in area 17 of Cebus monkey was conducted after iontophoretic injection of biocytin. Thirty axon terminals located in supragranular layers were qualitatively and quantitatively analyzed using 3-D automatic microscopy. Three types of axon terminals could be identified: Ia, Ib and II. Group I was characterized by a sparse and/or long-distance branch pattern, while type II presented compact and localized arborization. Ia axon terminals formed "clusters" and "terminaux"boutons while Ib did not. On overage, group II axon terminals tended to present straight or obtuse branching angles and a much more ramified pattern, and occupy a smaller cortical territory with shorter intermediate segments and higher density of synaptic potential sites than group I. The common characteristics of group I included innervation of larger cortical territories, longer intermediate segments, acute branching angles and lower synaptic density compared to group II. The results are compatible with the major subdivisions of neocortical neuronal morphology that classifies them as smooth and spine neurons. Smooth neurons may be related to axon terminals of group II while spine neurons may be related to group I.

Biocytin as a retrograde tracer in the mammalian visual system

We have successfully used biocytin as a retrograde tracer in the mammalian visual system. Retinal ganglion cells, pyramidal and stellate cortical neurons were labelled. Both pressure injections and gel implants were used successfully for retrograde labelling. Biocytin was detected using avidin conjugates and horseradish peroxidase histochemistry. Retrograde filling with biocytin proved to be more reliable and to allow better morphological resolution than other commonly used neurotracers such as horseradish peroxidase. The fine details of cell morphology observable by this method are comparable in many cases to the results obtained with intracellular tracer injections. The morphological resolution obtained with this method allows the study of brain microcircuits using extracellular deposits of biocytin

Head holder for lateral-eyed species in vision research

The present report describes a head holder designed to be used for lateral-eyed species in vision research. The head-holder employs a prosthesis implanted on the skull and provides for adjustable movements in different planes, thus allowing precise positioning of the eye with respect to the visual space