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Biomedical subjects

D Ehrlich

Publications and source records attributed to D Ehrlich.

At least 19 recordsLinked to original sources

Influence of acupuncture on physical performance capacity and haemodynamic parameters.

In a single blind study the question as to whether the needling of specific acupuncture points is able to produce an increase in physical performance capacity and better regulation of heart rate and blood pressure was examined. Thirty-six healthy young men were assigned at random to three groups, receiving either actual acupuncture, placebo acupuncture or no stimulation. Performance was determined by means of a spiro-ergometer test which was carried out at the beginning and at the end of five weeks of treatment consisting of one session per week. The subjects from the group which actually received acupuncture were able to increase maximum performance capacity significantly and also physical performance at the anaerobic threshold. This may be interpreted as a sign of functional improvement in haemodynamic and metabolic mechanisms. There was, on the whole, no noticeable effect produced by the placebo acupuncture. The control group, which received no stimulation, showed unfavourable changes in the values obtained compared with the results of the performance test at the commencement of the study.

Acupuncture Points

New thermosensitive plasmid for gram-positive bacteria.

We isolated a replication-thermosensitive mutant of the broad-host-range replicon pWV01. The mutant pVE6002 is fully thermosensitive above 35 degrees C in both gram-negative and gram-positive bacteria. Four clustered mutations were identified in the gene encoding the replication protein of pVE6002. The thermosensitive derivative of the related plasmid pE194 carries a mutation in the analogous region but not in the same position. Derivatives of the thermosensitive plasmid convenient for cloning purposes have been constructed. The low shut-off temperature of pVE6002 makes it a useful suicide vector for bacteria which are limited in their own temperature growth range. Using pVE6002 as the delivery vector for a transposon Tn10 derivative in Bacillus subtilis, we observed transposition frequencies of about 1%.

Amino Acid Sequence

Effects of selective neurotoxins on eye growth in the young chick.

We have determined the extent of retinal cell damage and eye growth after treatment with different neurotoxins. Day-old chicks received an intraocular injection containing 2, 10, 50, 100 or 200 nmol of kainic acid (KA), an excitotoxic analogue of glutamate. After 21 days, with 2 nmol KA damage was confined to a small proportion of bipolar cells, whereas with 10-200 nmol KA there was dose-dependent damage to amacrine cells, ganglion cells and photoreceptors. There was an increase in eye weight and size of the vitreal chamber with 10 nmol KA or more. In a similar protocol using 5, 50 or 200 nmol quisqualic acid (QUIS), there was massive loss of amacrine cells and slight loss of horizontal cells, and a large increase in the anterior chamber depth. A single dose of 200 nmol N-methyl-D-aspartate lesioned amacrine cells, but did not alter eye size. Thus, excitotoxins which have different retinotoxic effects also have different effects on eye size. In another study, we examined the effects of KA and QUIS on occlusion-induced eye enlargement. Treatment with 200 nmol QUIS alone resulted in increased depth of the anterior chamber. QUIS combined with occlusion for 3 weeks did not interfere with occlusion-induced growth of the vitreal chamber. Therefore, the loss of a large proportion of amacrine cells did not interfere with occlusion-induced growth. KA results in eye growth in its own right, but restricts occlusion-induced growth. These effects are associated with damage to photoreceptors, amacrine and bipolar cells and a small proportion of ganglion cells. Finally, we examined the effects of an intraocular injection of tunicamycin, a purported photoreceptor-specific neurotoxin in amphibia. Tunicamycin (0.1 or 1 mumg/eye) resulted in a flattened anterior chamber but did not affect growth of the vitreal chamber. Tunicamycin inhibited occlusion-induced growth, and treated retinae displayed massive disruption and loss of all cell types apart from a single row of pigmented retinal epithelium. We conclude that (a) normal and occlusion-induced eye growth may have separate mechanisms of action, (b) the vitreous and anterior chambers have different growth mechanisms, and (c) photoreceptors may play a critical role in occlusion-induced growth of the eye.

Animals

Differential effects of axotomy on substance P-containing and nicotinic acetylcholine receptor-containing retinal ganglion cells: time course of degeneration and effects of nerve growth factor.

The time course of degeneration of chick retinal ganglion cells was examined with Nissl stains and immunohistochemical methods for detection of substance P-like immunoreactive and nicotinic acetylcholine receptor immunoreactive neurons. Small lesions were made in the retinae, adjacent to the optic nerve head, and were subsequently sectioned parallel to the vitreal surface, permitting direct comparison of normal and axotomized retinal ganglion cells distal to the site of axon damage. At four and six days after surgery, a large number of degenerating cells with clear cytoplasm and pyknotic nuclei were seen. After eight, 10 and 14 days, many retinal ganglion cells displayed a chromatolytic response with dispersed Nissl granules, eccentric nuclei and the cells appeared crenulated. The number of apparently normal neurons in the ganglion cell layer in the axotomized region was reduced by about 50% six days following surgery, by about 70% on the 10th day and by about 75% on the 17th day. The remaining neurons in the ganglion cell layer were identified as displaced amacrine cells. From day 2 onwards, increased numbers of glial cells were present in the optic fibre, ganglion cell and inner plexiform layers. Many glial cells were enlarged and displayed extensive cytoplasmic processes, while others showed mitotic activity. Somata and proximal dendrites of retinal ganglion cells were intensely stained for substance P-like immunoreactivity at two and four days following surgery. At six, eight and 10 days, staining intensity was markedly reduced though still evident and at 14 and 17 days, substance P-like immunoreactivity had virtually disappeared. The persistence of limited substance P-like immunoreactive ganglion cells 10 days after surgery indicates that these cells have a relatively protracted response to axotomy. Nicotinic acetylcholine receptor-like immunoreactivity in the ganglion cells at two and four days following axotomy was substantially reduced. The majority of faintly stained nicotinic acetylcholine receptor-like immunoreactive ganglion cells, as visualized in counterstained sections, did not exhibit pyknosis in the immediate period following axotomy. Double label studies demonstrated that substance P-like immunoreactive ganglion cells were distinct from the nicotinic acetylcholine receptor-like immunoreactive ganglion cells. In a second set of experiments, nerve growth factor was then placed into the vitreous humor following intra-retinal axotomy. The somata, dendrites and proximal axons of lesioned substance P-like immunoreactive ganglion cells in these retinae were more intensely stained for a longer period of time and appeared more robust than cells from untreated retinae.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals

A quantitative analysis of the effects of excitatory neurotoxins on retinal ganglion cells in the chick.

The present study examines the differential effects of three excitotoxins, kainic acid (KA), N-methyl-D-aspartate (NMDA), and alpha-amino-2,3-dihydro-5-methyl-3-oxo-4-isoxazolepropanoic acid (AMPA) on neurons within the ganglion cell layer (GCL) of the chick retina. Two-day-old chicks were given a single, 5 microliters, intravitreal injection of KA, NMDA, or AMPA at a range of doses. Following treatment with 40 nmol KA, there was a 21% loss of neurons in the GCL. At 200 nmol KA, the loss increased to 46%. Exposure to KA eliminated mainly small neurons of soma area 5-15 microns2, and medium-sized ganglion cells of soma area 15-25 microns2. Large ganglion cells (greater than 25 microns2) remained unaffected. The vast majority of small cells were probably displaced amacrine cells. Exposure to 400 nmol NMDA resulted in a 14% loss of neurons, predominantly involving the large ganglion cells. At a dose of 3000 nmol NMDA, no further loss of cells was evident. Exposure to 200 nmol AMPA resulted in a 30% loss of large and some medium-sized ganglion cells. In a further series of experiments, exposure to excitotoxin was followed by a retinal scratch, which eliminated retinal ganglion cells within the axotomized region. The results indicate that only a small proportion of displaced amacrine cells are destroyed by NMDA and AMPA, whereas virtually all displaced amacrine cells are sensitive to KA. The findings of this study indicate the existence of subclasses of ganglion cells with specificity towards different types of excitatory amino acids (EAA).

Animals

Organisation of the hyperstriatal projection to the ventral lateral geniculate nucleus in the chick (Gallus gallus).

In the chick forebrain, the hyperstriatum accessorium (HA), receives an orderly ascending projection from the dorsolateral thalamus, a primary visual centre. The study examines the topography of the descending projection from HA to the ventral lateral geniculate nucleus (GLv), another retino recipient region of the diencephalon. Discrete injections of horseradish peroxidase-labelled wheat germ agglutinin were placed in the HA, and reaction product was assessed in terminals within the GLv. The rostrocaudal axis of HA undergoes a 180 degrees rotation and is represented along the caudorostral axis of GLv. A comparison with other inputs to GLv, retina and optic tectum, shows that all inputs are in visuotopic register.

Animals

Morphology of congenital microphthalmia in chicks (Gallus gallus).

This study examines the morphology of sporadic congenital microphthalmia in 1-day-old chicks, with particular emphasis on the neural retina. On the basis of the size of the eyeball it is possible to classify microphthalmia into two groups, severe and mild. In severe microphthalmia (less than 5 mm in equatorial diameter), the eyeball is severely malformed, but in most cases it shows evidence of an organized neural retina. Although ganglion cells and an optic nerve head are present in a small proportion of these retinae, we could not trace an optic nerve projection to the brain. These results indicate that some ganglion cells are able to be sustained after the period of naturally occurring cell death, suggesting either that those ganglion cells have established some contact with the central nervous system or that the presence of their axons in a rudimentary optic nerve is adequate for survival. In mild microphthalmia (greater than 5 mm in equatorial diameter), the most consistent abnormality is a defect in the pecten, which together with other abnormalities such as orbital cysts and colobomas indicates that the major abnormality occurs in the region of the choroid fissure. Associated with these defects are abnormalities within the ganglion cell layer. In some cases the number of ganglion cells was reduced, and in others the numbers of both ganglion and displaced amacrine cells were reduced. Unexpectedly, there were localized regions completely devoid of cells in the ganglion cell layer. The timing of the congenital defect may provide some clue as to the presence of a critical period in which displaced amacrine cells are formed or are sensitive to events related to ganglion cell loss.

Animals

Differential effects of excitatory amino acids on photoreceptors of the chick retina: an electron-microscopical study using the zinc-iodide-osmium technique.

Although excitotoxins derived from acidic amino acids are known to damage neurons in the inner nuclear and ganglion cell layers of the retina, little is known about their effects on photoreceptors. This study examines the acute and long-term effects of excitotoxins on photoreceptors of the chick retina. The zinc-iodide-osmium (ZIO) technique, which darkly labels a substantial subpopulation of synaptic vesicles in normal photoreceptor terminals, was used to supplement routine electron microscopy. Two-day-old chicks received a single intraocular injection of either 10, 50, or 200 nmoles kainic acid (KA), 200 nmoles N-methyl-D-aspartic acid (NMDA), or 200 nmoles quisqualic acid (QUIS), and were allowed to survive for either 6 h, 7 d, or 21 d. At 6 h, following exposure to 10, 50, and 200 nmoles KA, there was swelling and disruption of photoreceptor lamellae of the outer segments. At 7- and 21-d survival, 50 and 200 nmoles KA resulted in rounded, condensed synaptic terminals, which contained a high density of synaptic vesicles. However, there was complete loss of ZIO-positive vesicles within these photoreceptors. Outer segments were still disrupted, although small patches of lamellae were evident, suggestive of regeneration. Following exposure to QUIS, there was extensive swelling of outer segment lamellae at 6 h survival. Synaptic ribbons in terminals were also swollen. At longer survival periods, QUIS exposure resulted in a reduction of ZIO-positive vesicles, as well as swollen lamellae in outer segments. NMDA exposure, at either short or long-term survival, did not alter photoreceptor morphology, including the pattern of ZIO stain. The prolonged effects of KA, and to a lesser extent QUIS, on photoreceptors suggests that these drugs have a long-term effect on photoreceptor function. The ZIO technique provides a novel and potentially useful approach for identification of damaged photoreceptors.

Amino Acids

Excitatory amino acids interfere with normal eye growth in posthatch chick.

This study examines the effects of excitotoxic amino acids on eye growth and retinal morphology. Day old chicks received a single intraocular injection of either 200 nmoles kainic acid (KA), 200 nmoles quisqualic acid (QUIS) or 400 nmoles N-methyl-D,L-aspartate (NMDA). Following survival periods of 7, 14 and 21 days, eyeballs were removed and weighed. Measurements of axial length, equatorial length, anterior chamber depth and corneal diameter were taken. Treatment with KA increased eye weight and equatorial length. Treatment with QUIS increased the anterior chamber depth but decreased the equatorial length. Treatment with NMDA increased anterior chamber depth, but to a lesser extent than QUIS. The effects of QUIS and NMDA could be distinguished from those of KA since the former excitotoxins resulted in a marked increase in anterior chamber depth with no enlargement of vitreal chamber. Changes in eye size were evident by day 7 and were sustained throughout the duration of the experiment. Examination of retinae revealed that KA lesions amacrine cells, bipolar cells, some ganglion cells and photoreceptors. Exposure to QUIS lesions amacrine cells, horizontal cells and causes mild disruption of photoreceptor outer segments. In contrast, NMDA predominantly lesions amacrine cells. The results demonstrate that these neurotoxins have different effects on eye growth, which may be associated with differences in retinal pathology. It is proposed that photoreceptors are ideally suited to play a role in the control of eye growth.

Animals

Development of the supraoptic decussation in the chick (Gallus gallus).

The developing supraoptic decussation (SOD), a major interhemispheric tract in birds, has been implicated in both transfer of visual information and in the modulation of brain asymmetry. Moreover little is known of its morphology during development. We have examined the development of the chick SOD, which consists of three subregions; dorsal, ventral and subventral SOD. In the dorsal SOD the total number of fibres reach 968,000 on the 19th day of incubation (E-19), falling to 570,000 by the 8th day after hatching (P-8). In the ventral SOD, the number of fibres at E-19 reach 660,000, followed by a gradual reduction in their number to about 490,000 at P-22. In the subventral SOD the number of fibres estimated was 87,000 at E-15 falling to about 36,000 P-1. Compared with adult levels, there is, respectively, a drop in the number of fibres of 44%, 25% and 69% in the dorsal, ventral and subventral SOD during development. At E-19 in both the dorsal and ventral SOD there is qualitative evidence of axonal loss; disrupted axonal profiles, increased extracellular space and cells containing lysosomal cytoplasmic inclusions indicative of macrophages. Differences were also observed in the pattern of myelination, the dorsal, ventral and subventral SOD were shown to myelinate at different rates. Thus, in a single named tract, the SOD, there are major differences in the onset, rate and extent of fibre loss and myelogenesis within its three subregions. The functional implications of these differences are considered.

Animals

Neurotoxic effects of kainic acid on developing chick retina.

The neurotoxic effects of kainic acid (KA) on developing neurons in the chick retina was investigated in an in vitro preparation. Eyecups from chick embroyos at 6 (E-6), 8, 10, 12, 14, 16, and 20 days of incubation and from chicks on day 1 posthatch (D-1) were exposed to different doses of KA for 30 min and then processed for light microscopy. Neurotoxic damage was evaluated by the presence of swollen cell bodies, containing pale cytoplasm and pyknotic nuclei. At E-8, amacrine cells first became sensitive to KA and displayed neurotoxic damage at a threshold concentration of 20 microM. Their sensitivity to KA increased over the following 4 days, so that by E-12 they attained a threshold sensitivity of 5.0 microM KA. At E-14, one third of the retinae showed amacrine cell damage at 0.5-2.0 microM KA, less than the threshold dose of 5.0 microM KA required at D-1. Bipolar cells first become sensitive to KA at E-12, at a threshold concentration of 5.0 microM. The threshold concentration decreased over the following 10 days: 2.0 microM at E-16, 1.0 microM at E-20, and 0.5 microM at D-1. At E-8 and E-10, horizontal cells were susceptible to a relatively high concentration of 80 microM KA. The sensitivity to KA is evident prior to the formation of photoreceptor input. These results indicate that amacrine and horizontal cells are susceptible to KA at an earlier age than bipolar cells. Both amacrine and bipolar cells exhibit an age-dependent relationship with the threshold concentration of KA required to cause neurotoxicity; in general, the older the embryo, the lower the dose of KA. However, the increased susceptibility of amacrine cells at E-14 suggests a transient hypersensitivity to KA during this period which may reflect an overproduction of the receptor-ionic channel complex necessary for KA to exert its effect.

Animals

Distribution of substance P-like immunoreactive retinal ganglion cells and their pattern of termination in the optic tectum of chick (Gallus gallus).

Substance P-like immunoreactive (SP-LI) neurons were identified within the inner nuclear layer and ganglion cell layer of the chick retina. The SP-LI cells in the inner nuclear layer consisted of several subtypes of neurons, differing in soma size and dendritic arborization. In the ganglion cell layer a population of moderately labelled SP-LI neurons was also present. About 6-9 microns in diameter and spaced 50-80 microns apart, they formed a regular array across the entire retina, with a density of about 400 cells/mm2 in the superior temporal retina, declining to less than 100 cells/mm2 in the peripheral retina. The total number of SP-LI cells in the ganglion cell layer was approximately 75,000. Individual axons could be followed toward the optic nerve head. Lesions near the optic nerve head resulted in axotomy of ganglion cells within a limited portion of the retina. Two days of postaxotomy there were numerous SP-LI swellings in the proximal segments of axotomized axons. SP-LI neurons in the axotomized zone were larger, more numerous, and showed increased staining of their processes. Fourteen days following a retinal lesion, there was depletion of all SP-LI cells in the ganglion cell layer within the axotomized zone, but the SP-LI neurons in the inner nuclear layer were not noticeably affected. Following a localized injection of rhodamine-coupled latex beads into the optic tectum, a population of retinal ganglion cells (RGCs) in the contralateral retina was retrogradely labelled. Many of these cells also exhibited SP-like immunoreactivity. Examination of the optic tectum indicated the presence of SP-LI fibres in laminae 2-13 (nomenclature of Cajal: Histologie du Systeme Nerveux. Vol. 2. Paris: Maloine, '11), with immunoreactive terminal regions present mainly in laminae 2-4, 7, and 9-13. SP-LI cell bodies were found predominantly in laminae 10-12 and 13. Fourteen days following a retinal lesion, SP-LI processes and terminals were depleted from laminae 2 and 3. Immunoreactive cells and processes in the remaining laminae of the optic tectum were not noticeably altered. The present report confirms the existence of SP-LI retinal ganglion cells in the chick retina and demonstrates their contribution to lamina specific SP-LI arborization in the optic tectum.

Animals

Intravitreal kainic acid severely reduces the size of the developing optic tectum in newly hatched chickens.

Following a single intravitreal injection of 200 nmol of kainic acid (KA) to newly hatched chickens, there are acute and long-term effects on retinal ganglion cells in the chicken retina. Thirty min after injection, most ganglion cells showed cytoplasmic vacuolization. However, 14 days later, most ganglion cell soma appeared normal. Almost 60% of the cells in the ganglion cell layer (GCL) were lost, suggesting that displaced amacrine cells and not more than 40% of the ganglion cells had been eliminated. Following intravitreal injection of wheat germ agglutinin conjugated to horseradish peroxidase 14 days after the KA lesion, the amount of HRP reaction product was reduced in all retinorecipient layers, especially layers IIc and IId, of the tectum contralateral to the KA-treated eye. Fourteen days after the injection of kainic acid, during which the control tecta grow appreciably, all the superficial layers of the tectum contralateral to the kainic acid-lesioned eye, especially layers IIc and IId, were smaller than in controls, and did not differ in size from those seen in tecta contralateral to cut optic nerves. It is not clear whether this is a result of a developmental failure, or a shrinkage, or a combination of these factors. These results suggest that subtypes of ganglion cells may have a disproportionate influence in the maintenance of the cytoarchitectural integrity in the optic tectum. Alternatively the removal of the OFF-bipolar cells and amacrine cells presynaptic to ganglion cells may decrease their metabolism, and restrict the supply of trophic influences to the developing tectal cells.

Aging

Sequence analysis, cellular localization, and expression of a neuroretina adhesion and cell survival molecule.

A cDNA for purpurin, a secreted 20,000 dalton neural retina cell adhesion and survival protein, has been sequenced and expressed in mammalian cells. Purpurin mRNA is found in both embryonic and adult retina, but not the brain, heart, or liver. The protein is highly concentrated in the neural retina between the pigmented epithelium and the outer segments of the photoreceptor cells; it is synthesized by photoreceptor cells. The predicted purpurin sequence contains 196 residues, has approximately 50% sequence homology with serum retinol binding protein, and is a member of the alpha-2 mu-globulin superfamily. Purpurin binds retinol and may play a major role in retinol transport across the interphotoreceptor cell matrix.

Amino Acid Sequence

Specific ganglion cell death induced by intravitreal kainic acid in the chicken retina.

In young chickens, intravitreal kainic acid (60 nmol) causes axonal and terminal degeneration in some retinorecipient areas of the chicken brain. The accessory optic nuclei are markedly affected, suggesting that the large displaced ganglion cells are destroyed by kainic acid, and a specific pattern of degeneration is caused in the optic tectum, which suggests that other minor ganglion cell groups may also be sensitive to intravitreal kainic acid.

Animals

The avian pecten provides a potent substrate for growth and development of dissociated embryonic neural implants.

A cell suspension of the optic tecta of 3-day-old chick embryos was injected into the vitreal chamber of 2-day-old posthatch chicks. After a 14-21-day survival period, examination of eyeballs showed that all implants survived and, in 50% of cases, were attached to the pecten. The implants had proliferated and showed a laminated pattern of organization, with small cells in the superficial regions and large cells in the deep regions of the implant. The implants also contained a well-developed neuropil with mature synapses. The host retina was not affected by the presence of the implant. We suggest that the avian pecten represents a highly amenable structure for studies involving the response(s) by damaged retinae to neural implants.

Animals

Folic acid protects chick retinal neurons against the neurotoxic action of excitatory amino acids.

In this study we have examined the neurotoxic effects of folic acid (FA), alone or in combination with selected excitotoxins using in vitro preparations of chick retina. Folic acid alone at concentrations of up to 10 mM had no effect. Co-incubation of 10 mM FA with 2 microM kainic acid (KA) protected all cell types susceptible to KA toxicity, namely amacrine and bipolar cells. At lower concentrations the protective effect of FA to susceptible cell types was found to be dose-dependent. The rank order of cells which are protected by FA, in order of the lowest concentration of FA required, was amacrine, inner bipolar and outer bipolar cells. The effect of FA against KA is a very weak one, as a 130-400-fold concentration of FA is required to protect amacrine cells from KA and a 1000-5000-fold dose of FA required to protect bipolar cells. However, FA (2 mM) also protects susceptible retinal neurons from the neurotoxic effects of 40 microM N-methyl-DL-aspartic acid (NMDLA) and 60 microM quinolinic acid (QUIN); only requiring respectively 50 and 33 times the concentration of FA. Interestingly 10 mM FA had little effect against 40 microM quisqualic acid (QUIS). Thus FA antagonizes the effects of KA, NMDLA, QUIN and to a small extent QUIS. Although its action may be mediated through several receptor types, FA appears to be a more potent antagonist of the N-methyl-D-aspartic acid (NMDA)-preferring than the KA-preferring or QUIS-preferring receptor.

Animals

Retinal afferent and efferent connections in congenitally monophthalmic chicks.

The visual projections of the remaining eye of posthatch congenitally monophthalmic chicks were examined using wheat germ agglutinin-horseradish peroxidase. The morphology of the primary visual centres and their retinal projections contralateral to the injected eye were similar to those of normal chicks. The ipsilateral primary visual centres were smaller and less organized, yet all received retinal input. These ipsilateral retinal projections differ from those found in normal posthatch chicks [O'Leary et al.: Devl. Brain Res. 10: 93-109, 1983] in that they are more extensive and occupy some centres not previously reported to receive input. In the case of the ipsilateral isthmo-optic projection to the retina there was a substantial increase in the number of cells compared with normal chicks [O'Leary and Cowan: Devl. Brain Res. 12: 293-310, 1984]. A comparison of the extent of ipsilateral retinal afferents with that of normal chicks suggests that following loss of an eye two responses occur within the visual centres: in some centres there is a massive increase in the amount of ipsilateral terminals, whereas in others there is only a small increase. We propose that these responses are related to the intrinsic retinotopy within the visual centres. That is, highly retinotopic visual centres do not normally contain ipsilateral fibres, but following eye removal fibres from the ipsilateral eye are able to substantially innervate these regions. Presumably this effect is due to loss of the overriding influence of contralateral input, which would normally recognize and eliminate inappropriate ipsilateral fibres. In poorly retinotopic regions ipsilateral fibres are able to persist in both normal and monophthalmic chicks, as recognition cues may not be as precise as in highly retinotopic regions. Thus, the greater the retinotopic precision the finer the cues able to recognize ipsilateral fibres.

Animals