Modality-specific axonal projections in the CNS of the flies Phormia and Drosophila

There is a rich history of behavioral and physiological studies on the leg sensory systems of flies. Here we examine the anatomy of the sensory axons of two species of fly and demonstrate that the location of the axonal projections in the CNS can be correlated with the modality they encode. We studied receptors associated with proprioceptive, tactile, and multimodal hairs. Proprioceptive hairs occur in clusters, called hair plates, and are situated near joints. The neuron innervating each proprioceptive hair has a large axon and coarse arborization in the intermediate neuropil. Tactile receptors have smaller arbors, which are located in a ventral region of the thoracic neuromere. Finally, the multimodal hairs are each innervated by one tactile and four chemosensory neurons. The single tactile neuron has a central arbor that is indistinguishable from those of the tactile hairs; the four chemosensory neurons project to yet a third region of neuropil near the ventral surface of each neuromere. Thus there is a clear modality-specific segregation of axonal arbors in the CNS. This organization is identical in Phormia and Drosophila and thus apparently highly conserved within the Diptera. We presume that, as in other insect sensory systems, this anatomical specificity is linked to synaptic specificity.     

Position-specific central projections of mechanosensory neurons on the haltere of the blow fly,Calliphora vicina

The halteres of Dipteran insects play an important role in flight control. They are complex mechanosensory devices equipped with approximately 400 campaniform sensilla, cuticular strain gauges, which are organized into five fields at the base of each haltere. Despite the important role of these mechanosensory structures in flight, the central organization of the sensory afferents originating from the different field campaniforms has not been determined. We show here that in the blow fly, Calliphora vicina, sensory afferents from the campaniform fields project to the thorax in a region-specific manner. Afferents from different fields have different projection profiles and in addition, the projection pattern of afferents from different regions of the same field may show further variation. However, central target regions of these afferents are not exclusive to particular sensory fields because cells from different fields can possess similar projection profiles. Convergence of afferent projections is not limited to axons from the haltere fields, but is also observed between afferents originating from the haltere fields and those from serially homologous fields on the radial vein of the wing. Although we have not determined the specific cellular targets of the haltere sensory cells, the afferents of a dorsal field could make potential contact with at least one identified wing steering motoneuron that is known to be important in turning maneuvers. Our results, thus, provide the anatomical basis for studying how mechanosensory information encoded by the complex fields on the base of the haltere is mapped onto different functional regions within the CNS. © 1996 Wiley-Liss, Inc.     

Morphology and somatotopic organisation of the central projections of afferents from tactile hairs on the hind leg of the locust.

The morphology and organisation of the central projections of tactile hair afferents from the hind leg of the locust, Schistocerca gregaria, were examined by staining individual hair afferents. Each tactile hair on the femur, tibia, and tarsus is innervated by a single sensory afferent, which projects to the ipsilateral half of the metathoracic ganglion. Afferents arborize in the ventralmost and lateral ventral association centres (vVAC and lVAC). The projections are organised somatotopically in a map with three axes, according to the position of the hair on the leg. First, proximo-distal: afferents from hairs on the proximal leg segments project more anteriorly than do those from hairs on distal leg segments. Moreover, on any given segment the afferents from the more proximal hairs project more anteriorly than do the afferents from the distal hairs. Second, antero-posterior: afferents from hairs on the posterior surface of the leg project more medially than do afferents from anterior hairs. Third, dorso-ventral: afferents from hairs on ventral parts of the leg project more ventrally than do afferents from the dorsal hairs. The afferents from posterior and anterior hairs project to an area between the central projections from dorsal hairs and ventral hairs. The position of a projection within the map is dependent upon the location of the hair on the leg and not the peripheral routes taken by the axon of its afferent to reach the ganglion.     

Central neuronal projections and neuromuscular organization of the basal region of the shore crab leg

The musculature and associated skeleton, peripheral nervous system, and central projections of motor and sensory neurones of the two basal (thoracic and coxal) segments of the shore crab leg (fifth pereiopod, P5) were examined in vivo and with methylene blue or cobalt staining. Each of the four main basal muscles, promotor/remotor, levator/depressor, controlling the thoracico-coxal (T-C) and coxo-basal (C-B) limb joints, respectively, comprises several more or less discrete fibre bundles (total 14), with little morphological segregation of different functional groups. The innervation to the basal leg region is carried in two nerve roots arising from the thoracic ganglion. The anterior Th-Cx root carries both sensory and motor axons, while the posterior Th-Cx root is purely motor. Three previously undescribed sensory branches (two "epidermal" nerves and an "accessory" branch), in addition to that innervating the coxobasal chordotonal receptor, have been found in the distal part of the anterior Th-Cx root. Two clusters of 10 to 15 multipolar somata (diam. 30-125 micron) are located proximally at the bifurcation of the accessory nerve and distally where the latter enters the basipodite. The cell bodies (diameter 20-80 micron) of basal leg motoneurones (total ca. 30) lie in the dorsal cortex of the ganglion, with somata of functionally related motoneurones tending to form discrete structural groups. The morphology of individual motoneurones conforms to the general arthropod pattern. All are confined to the ipsilateral hemiganglion and their main neuropilar processes run parallel and in close apposition to each other with overlapping dendritic structures. Sensory projections arising from the CB chordotonal organ also ramify in the region of the neuropile invaded by motoneurones. The possible physiological significance of such structural associations within the CNS is discussed, as are the functional implications of basal limb anatomy in general.     

Innervation of dromyosuppressin (DMS) immunoreactive processes and effect of DMS and benzethonium chloride on the Phormia regina (Meigen) crop

Antibody to the dipteran myosuppressin peptide, dromyosuppressin, TDVDHVFLRFamide, stained cells and fibers in the brain, optic lobes, subesophageal ganglion, and thoracico-abdominal ganglion of the blow fly, Phormia regina (Meigen). Dromyosuppressin-like immunoreactive fibers were detected in the cardiac recurrent nerve, hypocerebral ganglion/corpora cardiaca complex, crop duct, and crop. In order to explore the mechanisms involved in regulating crop movement, we established an in vitro bioassay. The basal rate of crop movement was 50.8 +/- 1.5 contractions per minute. Application of 1 microl of saline to the crop did not significantly affect the rate of movement compared to the basal rate (46.1 +/- 1.1 contractions per minute, P < 0.05). Application of 1 microl 10(-6) M dromyosuppressin or 1 microl 10(-3) M benzethonium chloride to the crop slowed the rate to 2.2 +/- 0.2 and 6.1 +/- 0.7 contractions per minute, respectively. Although other data have previously been interpreted to suggest that dipteran crop contractions do not include a neural component, the neuropeptide dromyosuppressin affected P. regina crop motility. Innervation of the crop and crop duct by dromyosuppressin immunoreactive processes that originated in the central nervous system and the effect of dromyosuppressin on crop muscle contractions suggest that dromyosuppressin is released locally to modulate crop contractions and that crop motility is under neural regulation. Myosuppressins isolated from numerous insects have a high degree of structure identity and reduce spontaneous muscle contractions of the hindgut, oviduct, and heart. Benzethonium chloride, previously identified as a myosuppressin agonist on the cockroach hindgut and locust oviduct, mimicked the effect of dromyosuppressin on the crop. This suggests that structural requirements for myosuppressin receptor binding in the cockroach hindgut, locust oviduct, and fruit fly crop are similar.     

Central projections of coarse and fine vagal axons of the cat

We used the wallerian degeneration of vagal afferents and the retrograde transport of WGA-HRP microinjected in the nucleus of the tractus solitarius (NTS) to study the central projections of myelinated and unmyelinated vagal axons. We concluded that the set of largest nodose cells projected to the dorsolateral, interstitial, ventral, ventrolateral and intermediate NTS subnuclei, while the smaller nodose cells terminated in the medial, dorsal, gelatinosus and commissural NTS subnuclei.     

Specific expression of ezrin, a cytoskeletal-membrane linker protein, in a subset of chick retinotectal and sensory projections

Lamina-specific neuronal connections are a fundamental feature in many parts of the vertebrate central nervous system. In the chick, the optic tectum is the primary visual centre, and it has a multilaminated structure consisting of 15 laminae, of which only three or four receive retinal projections. Each of the retinorecipient laminae establishes synaptic connections selectively from one of a few subsets of retinal ganglion cells (RGCs). We have generated a series of monoclonal antibodies that appear to stain only one of the retinorecipient laminae. One of these, TB4, stained lamina F which receives inputs from a subpopulation of approximately 10-20% of RGCs which express the presynaptic acetylcholine receptor beta2-subunit. TB4 recognized a single 79-kDa protein on immunoblotting. cDNA cloning and immunochemical analysis revealed that the TB4 antigen molecule was ezrin, a cytoskeletal-membrane linker molecule belonging to the ezrin-radixin-moesin family. Unilateral enucleation of the eye, both prior to and after the establishment of retinotectal projections, attenuated the lamina-selective staining with TB4 in the contralateral tectum, suggesting that ezrin is anterogradely transported from RGCs to lamina F. Ezrin was thus expressed in a subset of RGCs that project to lamina F. Similar subset-selective expression and resultant lamina-selective distribution of ezrin were also observed in the lamina-specific central projections from the dorsal root ganglia. The staining pattern with TB4 in the dorsal root ganglia and spinal cord indicated that high expression of ezrin was restricted in cutaneous sensory neurons, but not in muscle sensory neurons. Since ezrin modulates cell morphology and cell adhesion profiles by linking membrane proteins with the cytoskeleton, it was suggested that ezrin is involved in the formation and/or maintenance of lamina-specific connections for neuronal subpopulations in the visual and somatosensory systems.     

From embryo to adult: anatomy and development of a leg sensory organ in Phormia regina, Meigen (Insecta: Diptera). II. Development and persistence of sensory neurons

The imaginal leg disc of Phormia regina contains eight neurons that arise during embryogenesis. Five of the neurons are associated with Keilin's organ, and of these five, two persist to the adult fly. Two new neurons arise at about the time of pupariation and flank each of these persisting neurons, forming two triplets of cells. Both triplets can be followed throughout metamorphosis; in the late pupa they are situated anteriorly and posteriorly at the tip of the fifth tarsomere. Two triplets of cuticular specializations are found at corresponding positions in the adult fly, each consisting of two campaniform sensilla and a trichoid hair. The central member of each set of sensilla, a campaniform sensillum, is associated with the persisting cell.     

Central course of digital axons within the median nerve of Macaca mulatta.

The traditional view that axons are not functionally grouped within proximal human nerve is based on the interfascicular dissections of Sunderland ('45). However, microstimulation and microneurography (Schady et al., '83a; Hallin, '90) reveal proximal grouping of cutaneous sensory axons from small areas of skin. In the present studies, conjugates of horseradish peroxidase with wheat germ agglutinin (HRP-WGA) were used to trace the course of digital nerve axons within the median nerve of Macaca mulatta. The electrophysiologic findings were confirmed, suggesting the potential for precise surgical realignment of functionally related axons even after proximal nerve transection. Radial digital nerves were labeled in the thumb (bilateral, 1 animal), the index finger (unilateral, 2 animals), and the middle finger (bilateral, 1 animal). Median nerve cross sections were cut at 1-cm intervals, treated with tetramethyl benzidine to demonstrate HRP-WGA within axons, and compiled to form maps of each digital nerve "territory" within the median nerve. These territories were limited to a single, densely labeled fascicle at the wrist level. They expanded somewhat in the forearm to encompass clusters of labeled axons within a matrix of unlabeled axon profiles. The clusters were more loosely packed in the arm, occupying 1/3 to 1/6 of the nerve cross section at the entrance to the brachial plexus. The three digital nerve territories studied were widely separated at the wrist level. In the proximal arm, there was moderate intermingling of axons from adjacent digits, but those to the middle finger and thumb remained segregated. Territory configuration differed widely overall, but was moderately constant for each digit. The location of territories within the nerve was often strikingly similar from right to left and from animal to animal, with occasional prominent variations reflecting isolated rotation of one nerve.     

The origin of postganglionic and sensory axons in the cervical sympathetic trunk of the cat: A horseradish peroxidase study

The composition of the cervical sympathetic trunk (CST) in the cat is still not completely understood. The present study investigates, by the horseradish peroxidase (HRP) method of tracing neuronal connections, the presence of postganglionic and sensory neurons projecting via the CST. Following sympathectomy at the midcervical level and the application of HRP crystals to the cut ends of the CST which had been isolated from the surrounding by a 1.5% solution of agar-agar, labelled neurons were seen in the superior cervical (SCG), stellate (SG), inferior vagal ganglia (IVG), and spinal ganglia C₈–T₈. The maximum number of labelled nuerons was 536 in the SCG, 460 in the SG, 180 in the IVG, and 104 in spinal ganglia C₈–T₈.     

Ultrastructure of the taste buds in the blind cave fish Astyanax jordani ("Anoptichthys") and the sighted river fish Astyanax mexicanus (Teleostei, Characidae)

This study describes the ultrastructure of the taste buds of the sighted river fish Astyanax mexicanus and of the blind cave fish Astyanax jordani (= Anoptichthys) (Teleostei, Characiformes, Characidae). In Astyanax and Anoptichthys, taste buds occur in the epithelia of the lips, oral cavity, and, in Anoptichthys, lower jaw. Both possess three types of taste buds: type I (elevated), type II (slightly elevated), and type III taste buds (not elevated or sunken). The taste buds are up to 60 microm high and up to 35 microm wide. The taste bud's sensory epithelium consists of 100--130 elongated cells: light cells, dense-cored-vesicles (dcv) -cells, dark cells, and degenerating cells. The dcv-cells are rich in dense-cored vesicles and are described for the first time in a teleostean taste bud. At the taste bud's base, there lie two to three basal cells. The basal cells of type I and type II taste buds have microvillus (spine)-like processes, in contrast to those of type III taste buds. The taste bud's nerve fiber plexus is situated between the bases of the elongated taste bud cells and the basal cells. Afferent synapses occur between dcv-cells and basal cells (presynaptic sides) and axons (postsynaptic side). Indistinct synapses occur between light cells and dark cells (presynaptic sides) and axons (postsynaptic side). The nerve fiber plexes of Anoptichthys type II and type III taste buds contain significantly more axon profiles than those of Astyanax. This may be associated with a compensatory improvement of the sense of taste in the blind, cave-dwelling fish.     

Two closely located areas in the suboesophageal ganglion and the tritocerebrum receive projections of gustatory receptor neurons located on the antennae and the proboscis in the moth Heliothis virescens

Sucrose stimulation of gustatory receptor neurons on the antennae, the tarsi, and the mouthparts elicits the proboscis extension reflex in many insect species, including lepidopterans. The sensory pathways involved in this reflex have only partly been investigated, and in hymenopterans only. The present paper concerns the pathways of the gustatory receptor neurons on the antennae and on the proboscis involved in the proboscis extension reflex in the moth Heliothis virescens (Lepidoptera; Noctuidae). Fluorescent dyes were applied to the contact chemosensilla, sensilla chaetica on the antennae, and sensilla styloconica on the proboscis, permitting tracing of the axons of the gustatory receptor neurons in the central nervous system. The stained axons showed projections from the two appendages in two closely located but distinct areas in the suboesophageal ganglion (SOG)/tritocerebrum. The projections of the antennal gustatory receptor neurons were located posterior-laterally to those from the proboscis. Electrophysiological recordings from the receptor neurons in s. chaetica during mechanical and chemical stimulation were performed, showing responses of one mechanosensory and of several gustatory receptor neurons. Separate neurons showed excitatory responses to sucrose and sinigrin. The effect of these two tastants on the proboscis extension reflex was tested by repeated stimulations with solutions of the two compounds. Whereas sucrose elicited extension in 100% of the individuals in all repetitions, sinigrin elicited extension in fewer individuals, a number that decreased with repeated stimulation.     

Ultrastructural analysis of NMDA,AMPA, and kainate receptors on unmyelinated and myelinated axons in the periphery

The present study determines the proportions of unmyelinated cutaneous axons at the dermal–epidermal junction in glabrous skin and of myelinated and unmyelinated axons in the sural and medial plantar nerves that immunostain for subunits of the ionotropic glutamate receptors. Approximately 20% of the unmyelinated cutaneous axon profiles at the dermal–epidermal junction immunostain for either N-methyl-D-aspartate (NMDA), α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA), or kainate receptor subunits. These findings are consistent with previous observations that NMDA and non-NMDA antagonists ameliorate nociceptive behaviors that result from noxious peripheral stimulation. In the sural nerve, where the large majority of myelinated fibers are sensory, approximately half of the myelinated axon profiles immunostain for the NMDA receptor 1 (R1) subunit, 28% immunostain for the glutamate receptor 1 (GluR1) AMPA subunit, and 11% for the GluR5,6,7 kainate subunits. Even higher proportions immunostain for these receptors in the medial plantar nerve, a mixed sensory and motor nerve. In the sural nerve, 20% of the unmyelinated axon profiles immunostain for NMDAR1 and only 7% label for GluR1 or GluR5,6,7. Because the sural nerve innervates hairy skin, these data suggest that glutamate will activate a higher proportion of unmyelinated axons in glabrous skin than in hairy skin. Measurements of fiber diameters indicate that all sizes of myelinated axon profiles, including Aδ and Aβ, are positively labeled for the ionotropic receptors. The presence of glutamate receptors on large-diameter myelinated axons suggests that these mechanosensitive receptors, presumably transducing touch and pressure, may also respond to local glutamate and thus be chemosensitive. J. Comp. Neurol. 391: 78–86, 1998. © 1998 Wiley-Liss, Inc.     

From embryo to adult: anatomy and development of a leg sensory organ in Phormia regina Meigen (Insecta: Diptera). I. Anatomy and physiology of a larval "leg" sensory organ

Neurons within the precursor of the adult leg, the imaginal disc, innervate a larval sense organ, Keilin's organ. Electron microscopical investigations of first instar larvae show that five dendrites end at the organ: three insert at the bases of the three hairs of the organ and two end against the cuticle, without any apparent cuticular specialization. In third instar larvae, the imaginal leg discs invaginate into the body cavity, and only four of the dendrites (the outer segments of which become greatly elongated) remain in contact with Keilin's organ. The axons of the neurons that supply Keilin's organ project into a ventral neuropile region of the central nervous system, with a pattern that resembles the projections of other larval sensilla. Electrical activity can be recorded from neurons of the imaginal disc in response to mechanical stimulation.     

Central projections from the skin of the hand in squirrel monkeys.

Central termination patterns of afferents from the hands of squirrel monkeys were studied after subdermal injections of wheat germ agglutinin conjugated with horseradish peroxidase (WGA-HRP) or cholera toxin subunit B conjugated to HRP (BHRP). WGA-HRP more effectively labeled axons terminating in the superficial dorsal horn of the spinal cord, while BHRP more effectively labeled axons terminating in the deeper layers. Injections of both tracers, when restricted to parts of glabrous digits, palm, or dorsal hand, revealed somatotopic patterns in the spinal cord and pars rotunda of the cuneate nucleus that were, in some respects, similar and, in other respects, quite different from those previously reported for macaque monkey (Florence et al., J. Comp. Neurol. 286:48-70, '89). As in macaques, injections in digits 1-5 produced a rostrocaudal sequence of foci of terminations in the cervical spinal cord. However, inputs from the palm were located medial to those from the digits, whereas the palm is represented lateral to the digits in macaque monkeys. Since inputs from the palm is also medial in the dorsal horn in cats (Nyberg and Blomqvist, J. Comp. Neurol. 242:28-39, '85), the condition in squirrel monkeys may be similar to the generalized state. In the cuneate nucleus, single injections in the hand produced dense label in the pars rotunda, and sparse label in the rostral and caudal poles. As in macaque monkeys, inputs from specific parts of the hand related to rostrocaudal clusters of cells that are cytochrome oxidase dense. The representation of the digits differed from macaques in that the digits were represented dorsal to the palm, rather that ventral to the palm as in macaques. Again, comparisons with cats suggest that squirrel monkeys have the more generalized pattern. Finally, inputs from the hair, dorsal surfaces of the digits terminated on the same clusters as the inputs from the glabrous, ventral surfaces, apparently overlapping somewhat. The proximity of these terminations from dorsal and ventral surfaces of the digits may be related to observations that cortical representations of the glabrous surfaces of digits become responsive to dorsal surfaces of the same digits when inputs from glabrous skin are chronically deactivated (e.g., Merzenich et al., Neuroscience 3:33-55, '83).     

Immunocytochemistry of gamma-aminobutyric acid, glutamate, serotonin, and histamine in Necturus taste buds.

Little information is currently available about which neurotransmitters are involved in signal processing in the peripheral sensory organs of taste, taste buds. Synaptic contacts between taste cells and sensory axons have long been known to exist, but what substances are active at these synapses is not known. Our objective in this study was to test for the presence of the neurotransmitter candidates, GABA, glutamate, serotonin, and histamine in taste buds of Necturus maculosus. Light microscopic immunocytochemical techniques were used to investigate the location of these substances in taste buds and surrounding epithelium. GABA and glutamate were detected in nerve fibers that innervate the taste buds, and, to a substantially lesser extent, in fine, varicose axons that penetrated the surrounding nontaste epithelium. Serotonin immunostaining was strong in basal cells in frog taste discs but was only faintly detected in Necturus taste buds. Histamine was not detected at all in taste buds. We conclude that amino acid neurotransmission may be involved in taste mechanisms and that monoamines may also play a role in chemosensory transduction in the taste bud. On the basis of our inability to detect histamine with immunocytochemical techniques, we conclude that this substance is unlikely to be a major neurotransmitter in Necturus taste buds.     

Distribution of choline acetyltransferase immunoreactive axons and terminals in the rat and ferret brainstem.

A survey was made of the density of the cholinergic innervation of different parts of the brainstem of the rat and ferret. Sections of rat and ferret brainstems were stained for choline acetyltransferase (ChAT) immunoreactivity by using a sensitive immunocytochemical method. Adjacent sections were stained for acetylcholinesterase activity or Nissl substance. The density of the distribution of fine calibre, varicose ChAT-positive axons, assumed to represent cholinergic terminals, was categorised arbitrarily into high, medium, or low. A high density of ChAT-positive terminals was found in all or parts of these structures: interpeduncular nucleus, superficial grey layer of the superior colliculus (ferret), intermediate layers of the superior colliculus, lateral part of the central grey (rat), an area medial to the parabigeminal nucleus (rat), pontine nuclei, ventral tegmental nucleus (rat), midline pontine reticular formation, and an area ventral to the exit point of the 5th nerve (ferret). A medium density of ChAT-positive terminals was observed in all or parts of: the substantia nigra zona compacta (ferret), ventral tegmental area (ferret), superficial grey layer of the superior colliculus, intermediate and deep layers of the superior colliculus, lateral central grey, area medial to the parabigeminal nucleus, inferior colliculus, dorsal tegmental nucleus, ventral tegmental nucleus (ferret), pontine nuclei, ventral nucleus of the lateral lemniscus (ferret), midline pontine reticular formation, ventral cochlear nucleus, dorsal cochlear nucleus, lateral superior olive, spinal trigeminal nuclei, prepositus hypoglossal nucleus, lateral reticular nucleus, paragigantocellular nucleus, and the dorsal column nuclei including the cuneate, external cuneate, and gracile nuclei. A low density of ChAT-positive terminals was seen throughout the remainder of the brainstem of the rat and ferret, but these terminals were absent from the medial superior olive, substantia nigra zona reticulata (rat), and the central part of the ferret lateral superior olive. A pericellular-like distribution of ChAT-positive terminals was observed in the ventral cochlear nucleus and in association with some of the cells of the nucleus of the mesencephalic tract of the trigeminal nerve. A climbing fibre type arrangement of ChAT-positive terminals was found in the substantia nigra zona compacta (ferret) and medial reticular formation. In general, the distribution of staining for AChE activity reflected that of the distribution of ChAT immunoreactivity in the brainstem, except in a few regions where there were also species differences in the distribution of ChAT-positive terminals, e.g., in the superficial grey layer of the superior colliculus and in the substantia nigra.(ABSTRACT TRUNCATED AT 400 WORDS)     

Position of axons in the cat's optic tract in relation to their retinal origin and chiasmatic pathway.

The positions of the crossed and uncrossed optic axons of distinct diameter classes has been examined in the optic tract of the adult cat. In addition, the retinal origin of axons occupying different positions within the tract has been studied. Since the position of a fibre within the optic tract reflects its time of arrival during development, we have used axonal position as an indicator of age and have related this to the chiasmatic pathway choice of the axons. Cats were either monocularly enucleated, to reveal the position and diameter of surviving crossed and uncrossed optic axons in semithin and thin sections, or implants of horseradish peroxidase (HRP) were placed so as to retrogradely label the ganglion cells giving rise to axons within the deep (early arriving), or superficial (later arriving) parts of the tract selectively. This was accomplished by either 1) surgically implanting HRP into the superficial portion of the optic tract, via a transbuccal approach, or 2) making such a transbuccal transection of the superficial fibres, followed by intracerebral injections of HRP to retrogradely label the surviving, deeper, optic axons from their target nuclei. The deep parts of the optic tract contain fine and medium, crossed and uncrossed axons arising from mainly medium sized cells in the contralateral nasal and the ipsilateral temporal retina; there is a clear line of decussation. In contrast, the superficial parts of the tract contain mainly fine diameter axons arising from small cells in the whole contralateral retina, and a small proportion of large diameter axons arising from large, alpha cells in the whole contralateral retina and in the ipsilateral temporal retina. The likelihood that axons from the temporal retina will project contralaterally therefore increases as development proceeds, since these axons are found in the superficial parts of the tract only. This suggests that a time-dependent signal that weakens with age is responsible for directing early arriving optic axons from the temporal retina to take an exclusively uncrossed course.