Distribution of omega-Conotoxin GVIA binding sites in teleost cerebellar and electrosensory neurons.

The distribution of omega-Conotoxin GVIA (CgTx) binding sites was used to localize putative N-type Ca2+ channels in an electrosensory cerebellar lobule, the eminentia granularis pars posterior, and in the electrosensory lateral line lobe of a gymnotiform teleost (Apteronotus leptorhynchus). The binding sites for CgTx revealed by an anti-CgTx antibody had a consistent distribution on somatic and dendritic membranes of specific cell types in both structures. The distribution of CgTx binding was unaffected by co-incubation with nifedipine or AgaToxin IVA, blocking agents for L- and P-type Ca2+ channels, respectively. Incubation with CgTx in the presence of varying levels of extracellular Ca2+ altered the number but not the cell types exhibiting immunolabel. A punctate immunolabel was detected on somatic membranes of granule and stellate cell interneurons in both the eminentia granularis pars posterior and the electrosensory lateral line lobe. Punctate CgTx binding sites were also present on spherical cell somata and on the large presynaptic terminals of primary afferents that terminate on spherical cells in the electrosensory lateral line lobe. No label was detected in association with distal dendritic membranes of any cell class, or with parallel fibers in the respective molecular layers. Binding sites for CgTx in the eminentia granularis are consistent with the established role for N-type Ca2+ channels in cell migrations, an activity which is known to persist in this layer in adult Apteronotus. The distribution of labeled stellate cells with respect to topographic maps in the electrosensory lateral line lobe further suggest that N-type Ca2+ channels are expressed in relation to functional activity across these sensory maps.

Nitric oxide synthase in tiger salamander retina.

Previous studies have indicated that nitric oxide, a labile freely diffusible biological messenger synthesized by nitric oxide synthase, may modulate light transduction and signal transmission in the retina. In the present work, the large size of retinal cells in tiger salamander (Ambystoma tigrinum) allowed the utilization of nicotinamide adenine dinucleotide phosphate (NADPH)-diaphorase histochemistry and nitric oxide synthase immunocytochemistry to delineate the cell-specific intracellular localization of nitric oxide synthase. NADPH-diaphorase activity was highly concentrated in the outer retina, in rod and cone inner segment ellipsoids, and between and adjacent to the photoreceptor cell bodies in the outer nuclear layer. Examination of enzymatically isolated retinal cells indicated that outer nuclear layer NADPH-diaphorase activity was localized to the distal processes of the retinal glial (Müller) cells and to putative bipolar cell Landolt clubs. Less intense NADPH-diaphorase activity was seen in the photoreceptor inner segment myoid region, in a small number of inner nuclear layer cells, in cap-like configurations at the distal poles of cells in the ganglion cell layer and surrounding ganglion cell layer somata, and in punctate form within both plexiform layers, the pigment epithelium, and the optic nerve. Nitric oxide synthase-like immunoreactivity was similarly localized, but was also concentrated along a thin sublamina centered within the inner plexiform layer. The potential for nitric oxide generation at multiple retinal sites suggests that this molecule may play a number of roles in the processing of visual information in the retina.

ON and OFF activity gradients in the lateral geniculate nucleus of the cat: a combined 14C 2-deoxyglucose and D,L-2-amino-4-phosphonobutyric acid study.

Experiments on the lateral geniculate nucleus (LGN) of the cat based on 14C 2-deoxyglucose (2-DG) autoradiography and intraocular injections of 2-amino-4-phosphonobutyric acid (APB) provided evidence for gradients of metabolic activity in the ON and OFF pathways in layer A, but only very weakly, if at all, in layer A1. Alert and freely moving cats were exposed to square-wave gratings over a 45-min period after injection of the 2-DG. When one eye had been treated previously with APB, contralateral layer A showed a clear gradient of 2-DG label indicating that the remaining OFF pathway was most active ventrally in the layer and, by implication, that the ON pathway is normally most active dorsally. No gradient was apparent in layer A1 ipsilateral to the APB eye. Control experiments based on binocular injections of tetrodotoxin (TTX) demonstrated that no gradients were present in the baseline activity within the layers. Finally, monocular injections of TTX provided evidence for gradients of nondominant eye activity in layers A and A1 that were maximal near the interlaminar zone between layers A and A1 and declined in mirror-symmetric fashion toward the dorsal border of A and the ventral border of A1. Combined with earlier anatomical studies showing depth-dependent patterns of geniculo-cortical projection, these results indicate that in the cat, as in several other species, the visual input to striate cortex is partly organized around ON and OFF pathways. In addition, the results suggest that a systematic variation of binocular interaction, perhaps related to ocular dominance, exists through the depths of the geniculate layers.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of flashing-diffuse light on [2-14C]deoxyglucose uptake in the visual system of the black-hooded rat.

What components of the visual system process diffuse light information? A [2-14C]deoxyglucose (2-DG) autoradiographic analysis revealed that exposure of freely moving rats (wearing light-diffusing masks) to flashing-diffuse light consistently elevated 2-DG uptake in the lateral geniculate nucleus and superior colliculus to levels rivalling those occurring in rats exposed to flashing-gratings. Uptake in visual cortex (area 17) in response to flashing-diffuse light, however, varied as a function of early contour experience, i.e. lower than that produced by darkness in rats reared with high contrast patterns, higher than darkness in rats which had been lid-sutured from the time of eye opening, and falling between these two extremes in 'ordinary' cage-reared rats. The findings point to subcortical mediation of discriminations based on diffuse light information. Cortex might participate in the processing of diffuse light information in the special case of animals lacking contour experience during development.

[2- 14C]deoxyglucose uptake in rat visual system during flashing-diffuse and flashing-pattern stimulation over a 6 log range of luminance.

The 2-deoxyglucose autoradiographic technique was used to assess the metabolic activity of cortical area 17, the dorsal and ventral lateral geniculate nuclei, the lateral posterior nucleus, and the superior colliculus, during 5-Hz flashing-pattern (montage of black and white square-wave gratings) and flashing-diffuse (eye covered with white mask) stimulation at three intensities over a 6 log range. In area 17 flashing-pattern was found to be equally effective at elevating uptake of the functional label over the photopic-scotopic range of luminance levels tested, whereas flashing-diffuse was ineffective. In subcortical nuclei, however, flashing-diffuse was no less effective than flashing-pattern and uptake of the label correlated positively with intensity level. The results suggest that the subcortical components of the visual system do play an important role in the processing of intensity information and that primary visual cortex does not.

Activity-dependent changes in eye influence during monocular blockade: increases in the effects of visual stimulation on 2-DG uptake in the adult rat geniculostriate system.

We examined the effects of loss of monocular retinal activity on 2-deoxyglucose (2-DG) uptake in the adult rat geniculostriate system. Of particular interest was whether the influence of the normally functioning eye changed during long-term contralateral retinal silence. Group 1 rats were subjected to short-term (24 hours) and group 2 rats to long-term (21-90 days) monocular tetrodotoxin (TTX) blockade, and metabolic activity was assessed during exposure to square-wave gratings. Group 1 rats exhibited patterns of cortical glucose utilization commensurate with complete monocular loss of retinal activity: minimal 2-DG uptake in contralateral monocular area 17 and dorsal lateral geniculate nucleus (LGN), and a bilateral depression in the binocular regions; 2-DG uptake was highest in the monocular regions fed by the stimulated normal eye (in both area 17 and the LGN) and these regions appeared unaffected by the monocular blockade. After repeated injections of TTX (group 2), metabolic activity in binocular area 17 and binocular LGN increased bilaterally relative to the metabolically active monocular regions contralateral to the normal eye. Group 3 rats were monocularly TTX-injected for 30 or 60 days, and, 24 hours before 2-DG, all retinal activity was eliminated by means of binocular TTX injections or binocular enucleation. Glucose utilization in the binocular regions of both area 17 and the LGN in these rats was depressed to levels seen in monocular area 17 after complete and recent loss of activity from the contralateral eye, indicating that the metabolic increase which occurred in the binocular regions during long-term monocular retinal blockade was dependent upon the neuronal processing of retinal information from the non-TTX eye. We conclude that, in the adult rat, an activity-dependent, physiologically based shift in ocular influence occurred in the binocular geniculostriate system during long-term monocular retinal inactivation.

Effects of prolonged retinal ganglion cell inactivity on superior colliculus glucose metabolism in the mature hooded rat.

Previous work utilizing the 2-deoxyglucose (2-DG) technique demonstrated that denervation, both direct (enucleation) or indirect (retinal receptor destruction, leaving ganglion cells intact), resulted in a depression followed by an increase in glucose metabolism in the superior colliculus (SC) of the mature hooded rat. Both enucleation and receptor loss result in (1) cessation of ganglion cell activity and (2) disruption of connections between visual system neurons. To examine the relative importance of these two factors to the metabolic depression-"recovery" sequence, retinal ganglion cells were silenced without denervation for periods ranging from 24 h to 2 months by means of repeated intraocular tetrodotoxin (TTX) injections. SC metabolic activity fell to levels comparable to those seen after enucleation or receptor damage, but no recovery was detected. A more sensitive within-animal comparison was carried out to detect any small shifts in metabolic activity which might have occurred during retinal blockade; after 1 or 2 months of monocular TTX treatment, either binocular enucleation or binocular TTX injections were performed 24 h before 2-DG, depriving both sides of the SC of retinal input. Metabolic activity was slightly higher in the SC that had received no retinal input for 1 or 2 months, indicating that physiological changes had occurred within the SC during the afferent blockade. A second group of rats was allowed to recover from the effects of long-term TTX for from 1 to 12 days and binocularly exposed to visual stimuli during 2-DG uptake to determine whether damage had resulted from the TTX injections and to assess the time course of the effects of retinal blockade on SC functional activity.(ABSTRACT TRUNCATED AT 250 WORDS)

Metabolic activity in striate and extrastriate cortex in the hooded rat: contralateral and ipsilateral eye input.

The extent of changes in glucose metabolism resulting from ipsilateral and contralateral eye activity in the posterior cortex of the hooded rat was demonstrated by means of the C-14 2-deoxyglucose autoradiographic technique. By stimulating one eye with square wave gratings and eliminating efferent activation from the other by means of enucleation or intraocular TTX injection, differences between ipsilaterally and contralaterally based visual activity in the two hemispheres were maximized. Carbon-14 levels in layer IV of autoradiographs of coronal sections were measured and combined across sections to form right and left matrices of posterior cortex metabolic activity. A difference matrix, formed by subtracting the metabolic activity matrix of cortex contralateral to the stimulated eye from the ipsilateral "depressed" matrix, emphasized those parts of the visual cortex that received monocular visual input. The demarcation of striate cortex by means of cholinesterase stain and the examination of autoradiographs from sections cut tangential to the cortical surface aided in the interpretation of the difference matrices. In striate cortex, differences were maximal in the medial monocular portion, and the lateral or binocular portion was shown to be divided metabolically into a far lateral contralaterally dominant strip along the cortical representation of the vertical meridian, and a more medial region of patches of more or less contralaterally dominant binocular input. Lateral peristriate differences were less than those of striate cortex, and regions of greater and lesser monocular input could be distinguished. We did not detect differences between the two hemispheres in either anterior or medial peristriate areas, thus indicating either completely binocular input (which seems unlikely given the retinotopic organization of these regions), or a greater dependence than in the lateral peristriate on inputs that were not affected by the visual manipulations.

Organization and properties of neurons in a visual area within the insular cortex of the cat.

1. Extracellular recordings from 304 neurons were obtained with carbon fiber-containing multibarrel micropipettes. The cells were isolated in the insula in cats anesthetized with barbiturate and immobilized with gallamine triethiodide. Cells were tested with visual stimuli in the form of bars of light, moving edges, and square-wave, drifting grating patterns. 2. The spatial extent of the visually responsive region of insular cortex was assessed and was found to be limited to a surface area of approximately 6-8 mm2, the perimeter being delimited caudally by visually unresponsive cortex of the anterior sylvian gyrus, rostrally by the cortex surrounding the posterior third of the orbital sulcus (ventral bank), dorsally by the rostral extension of the dorsal bank of the anterior ectosylvian sulcus, and ventrally by a visually unresponsive zone bounded by a region about 2 mm2 ventrolateral from the anterior ectosylvian sulcal infolding. Furthermore, a group of unimodal, visually responsive cells often was found in the upper bank of the anterior rhinal sulcus. 3. The possibility of there being a visuotopic organization of insular neurons was examined by analyzing the distribution of receptive-field representation of neurons in sequential penetrations, as well as by searching for spatial progressions in the locations of visually responsive areas within the region. No such clear-cut organization was found among the cells of the insula. 4. Visually responsive neurons were encountered in groups, within electrode penetrations. These groupings were roughly segregated into three distinct levels within the depth of the cortex: the first between the pial surface and 600 micron, the second between 1,100 micron and 1,800 micron, and the third between 2,000 micron and 2,500 micron. 5. Neurons were classified according to their velocity sensitivity, directional preference, orientation sensitivity, length preference, modality specificity, response to electrical stimulation of extrageniculostriate regions, and response to light stimulation in the presence of microiontophoretically administered bicuculline methiodide (BMI). 6. Cells of superficial layers tended to exhibit a preference for high-velocity movements of light bars (600 degrees s-1), whereas those of deeper laminae generally preferred relatively lower velocity movements (60 degrees s-1). The clear preferences of many cells for certain directions of movement within the 360 degrees arc suggested the presence of a dynamic orientation sensitivity. 7. Proportionately more cells preferred moving bars (57%) to small moving spots (43%).(ABSTRACT TRUNCATED AT 400 WORDS)

Modality specificity of neuronal responses within the cat's insula.

1. Electrophysiological recordings of single-unit responses, multiunit responses, and electrically evoked field potentials have been made using carbon fiber-containing micropipettes in cats anesthetized with barbiturate and immobilized with gallamine triethiodide. Recording sites sampled cortical regions throughout the insula, including zones more ventrally situated and more rostral and caudal than those described in the preceding, companion paper. One-hundred eleven cells in total were tested with a battery of different types of stimuli. 2. Stimuli were divided into two classes, according to either the intensity of the stimulus or its form. These are called physiological forms, or levels of stimulation, and nonphysiological forms or levels. The nonphysiological forms of stimuli for visual, somatosensory, and auditory modalities consisted of (for visual stimuli): 1) electrical stimulation of the optic nerve or 2) bright flashes light at 100% contrast; for somatosensory, electrical stimulation of the radial nerve by implanted cuff electrodes; and for auditory, stimulation with bursts of white noise generated at high intensities (80-100 dB) or with a loud click stimulus. Physiologically relevant levels of stimuli for these same modalities were: moving bars of light projected onto a tangent screen in front of the animal (visual); light cutaneous deformation, hair displacement, and light pressure delivered to various regions on the surface of the cat's body with hand-held probes, or delivered manually (somatosensory); and white noise generated at low intensities (ca. 40 dB) (auditory). 3. Cells situated in dorsal insular regions responded to visual stimuli when levels of sensory activation were employed using natural means, within normal, physiologically relevant limits. Responses to auditory or somatosensory stimulation were observed in this region only when very intense forms of "natural" stimulation, or when electrical stimulation (nonphysiologically relevant levels of stimulation) was delivered. In this latter case, the same cells in several instances could be made to appear polymodally responsive. With cells situated in ventral insular regions, some polymodal responses to physiologically relevant levels of stimulation were noted, although it was considerably more common to obtain unimodal responses. Nonphysiological levels of activation yielded evidence for a polymodal convergence onto the greater proportion of cells recorded. 4. Field potential recordings with microelectrodes revealed widely overlapping representations of all modalities in both dorsal and ventral regions of the insula, irrespective of the sensitivity displayed by the local neuronal r

Metabolic changes in the superior colliculus after retinal receptor loss--neurotrophic interactions in the inactive visual system.

In the mature rat, direct denervation by means of eye enucleation resulted in a temporary metabolic depression followed by "recovery" in primary visual centers as determined by the 2-deoxyglucose technique (4). After unilateral destruction of the retinal receptor layer by means of intense light, the superior colliculus (SC) demonstrated this same depression-recovery process. Because receptor destruction is believed to silence ongoing ganglion cell activity, and because the SC changes occurred whether or not ganglion cells sustained damage, it appeared that direct denervation of colliculus neurons was not necessary to initiate the depression-recovery sequence and that lack of activity or "disuse" was the critical factor. The silencing effect of the receptor destruction was confirmed when tetrodotoxin (TTX) injections into the damaged eye 2 months after damaging light exposure only slightly affected metabolic activity in the recovered colliculus. Binocular TTX injections in unilaterally light-damaged rats after 2 months of recovery resulted in greater depression in the normal colliculus than in the "recovered" colliculus, again suggesting that increases in glucose metabolism over time reflected physiological adjustments in the SC to loss of afferent activity. The strong depression in the SC fed by the normal eye after TTX injection confirmed that tonic retinal afferent activity was important to the metabolic integrity of the SC and that cessation of such activity could lead to at least to depression in the system. In a final group of 2-month recovery animals the light-damaged eye was enucleated. Presumably, if withdrawal of afferent activity is solely responsible for initiating the depression-recovery sequence, the destruction of already silenced retinal ganglion cells would have no effect on the recovered SC. This was not found to be the case. In fact, enucleation reinstated the metabolic depression in the recovered SC and demonstrated that denervation per se resulted in depression of glucose metabolism in postsynaptic neurons. Even in the absence of impulse activity, visual system neurons maintained trophic interactions.

Reversal of 2-deoxyglucose uptake pattern across rat superior colliculus layers during recovery from visual cortex ablation.

The 2-deoxy-D-[14C]glucose autoradiographic technique was used to assess metabolic activity across stratum griseum superficiale, stratum opticum, and stratum griseum mediale of the superior colliculus, at intervals of 1 to 90 days after a unilateral visual cortex lesion. Initially, glucose metabolism ipsilateral to the lesion was more depressed in stratum griseum mediale than in superficiale, but beginning at about 14 days postlesion this layer pattern reversed. The rise in mediale metabolic activity correlated in time with a decline in generalized cortical depression. On the basis of anatomic connections, we concluded that the generalized cortical depression had disrupted input to stratum griseum mediale from cortical areas other than area 17, and that with recovery of cortical metabolic activity, stratum griseum mediale metabolic activity increased to nearly normal.

Increased dependence of superior colliculus metabolic activity on visual cortex after eye enucleation.

Metabolic activity, as previously shown by the 2-deoxy-D-[14C]glucose technique, is depressed in the superior colliculus after eye enucleation and recovers substantially by 30 days. To determine whether or not this recovery involves an increase in visual cortex control over superior colliculus metabolic activity, rats that had undergone either monocular or binocular enucleation 30 days earlier received visual cortex ablations. In the monocularly enucleated group, a bilateral visual cortex lesion produced greater depression in the recovered superior colliculus than in the opposite control colliculus, and increased the metabolic differences between the two colliculi compared with those of rats recovered from monocular enucleation but cortically intact. In the binocularly enucleated group a unilateral cortex lesion produced greater ipsilateral depression than did the same ablation in the otherwise intact rat. These findings led to the conclusion that the recovery in the superior colliculus that follows eye enucleation involved an increase in cortical control over metabolic activity.

Depression and recovery of metabolic activity in rat visual system after eye removal.

To examine the anterograde metabolic effects of visual system damage, unilateral eye enucleation was carried out in 19 black-hooded rats, and the animals were injected with 2-deoxy[14C]glucose at postoperative survival times ranging from 1 h to 91 days. Metabolic depression followed by recovery to near-normal resting levels of activity was seen contralateral to the enucleation in the superior colliculus and the dorsal and ventral lateral geniculate nucleus (primary effects), and in the lateral posterior nucleus, in layer IV, and the infragranular layers of visual cortex (secondary effects). Diaschisis, defined as a temporary depression in neural activity after denervation, appears to be a characteristic response of the damaged visual system.

2-Deoxyglucose uptake in the thalamus of awake rats after neocortical ablations.

Local, temporary or permanent depressions and elevations in 2-deoxy[14C]glucose (2-DG) uptake were observed in the brains of unanesthetized, freely moving rats that 3 days to 1 year earlier had sustained large unilateral posterior neocortical ablations. Some of the differences in metabolic activity between the normal and damaged hemispheres, in both cortical and subcortical regions, were attributed to cell death and gliosis, and surgical trauma. In addition, it appeared that certain metabolic changes provided a physiologic correlate of diaschisis, a temporary impairment in function which von Monakow hypothesized to occur in neurons denervated by a lesion. We concluded that the 2-DG technique is a useful tool for examining the widespread effects of brain damage, and that when employed in the manner exemplified by this and a previous study, the technique can disentangle the changes in neural activity which occur after brain damage from the changes in glial activity which accompany them.

2-Deoxyglucose uptake and histologic changes in rat thalamus after neocortical ablations.

2-Deoxy[14C]glucose (2-DG) uptake was examined in pentobarbital-anesthetized rats that 3 days to 2 years earlier had sustained large posterior neocortical ablations. At 2 weeks postoperatively 2-DG uptake was pronounced in those thalamic nuclei with cortical projection zones included in the ablated region. At shorter postoperative intervals (3 to 4 days) there appeared to be no increases in 2-DG uptake in these thalamic nuclei and at longer postoperative intervals (greater than 1 month) increases were only discernible in calcified sites. Both temporal and spatial postoperative variations in 2-DG uptake were found to coincide almost perfectly with glial cell proliferation and staining in the affected regions.