Central pupillary light reflex circuits in the cat: II. Morphology,ultrastructure, and inputs of preganglionic motoneurons

Preganglionic motoneurons supplying the ciliary ganglion control lens accommodation and pupil diameter. In cats, these motoneurons make up the preganglionic Edinger‐Westphal population, which lies rostral, dorsal, and ventral to the oculomotor nucleus. A recent cat study suggested that caudal motoneurons control the lens and rostral motoneurons control the pupil. This led us to examine the morphology, ultrastructure, and pretectal inputs of these populations. Preganglionic motoneurons retrogradely labeled by introducing tracer into the cat ciliary ganglion generally fell into two morphologic categories. Fusiform neurons were located rostrally, in the anteromedian nucleus and between the oculomotor nuclei. Multipolar neurons were found caudally, dorsal and ventral to the oculomotor nucleus. The dendrites of preganglionic motoneurons within the anteromedian nucleus crossed the midline, providing a possible basis for consensual responses. Ultrastructurally, several different classes of synaptic profiles contact preganglionic motoneurons, suggesting that their activity may be modified by a variety of inputs. Furthermore, there were differences in the synaptic populations contacting the rostral vs. caudal populations, supporting the contention that these populations display functional differences. Anterogradely labeled pretectal terminals were observed in close association with labeled preganglionic motoneurons, particularly in the rostral population. Ultrastructural analysis revealed that these terminals, packed with clear, spherical vesicles, made asymmetric synaptic contacts onto motoneurons in the rostral population, indicating that these cells serve the pupillary light reflex. Thus, the preganglionic motoneurons found in the cat display morphologic, ultrastructural, and connectional differences suggesting that this rostral preganglionic population is specialized for pupil control, whereas more caudal elements control the lens. J. Comp. Neurol. 522:3978–4002, 2014. © 2014 Wiley Periodicals, Inc.     

Cells in the pretectal olivary nucleus are in the pathway for the direct light reflex of the pupil in the rat

Extracellular microelectrode recordings from 148 single cells in the pretectum of the hooded rat were classified according to their temporal response properties to light stimulation of their retinal receptive fields. Fifty-six cells were classified as tonic-on cells, 22 cells were classified as tonic-off cells, and 53 cells were classified as phasic cells. Seventeen cells could not be assigned to one of these 3 groups. The diameters of the receptive field centers of the tonic-on pretectal cell were clustered about a mean of 31° and the temporal response of these cells was sustained. Constriction of the contralateral pupil was produced by electrical stimulation through the recording electrode at sites containing tonic-on pretectal cells, but not at sites containing tonic-off pretectal cells or phasic pretectal cells. For this reason, we argue that tonic-on cells are likely to mediate constriction in the light reflex of the rat's pupil. Receptive field maps together with electrolytic marking lesions at recording and stimulation sites showed that tonic-on pretectal cells are retinotopically organized and are aggregated in a strip running from the dorso-medial tip of the pretectum to the ventro-lateral boundary. The anatomical distribution of these cells is coextensive with the region known as the pretectal olivary nucleus (PO) in the rat^26,27.Using fine microelectrodes, recordings were obtained from 27 axons presumed origin (fibers). Of these, 14 were tonic-on, 10 were phasic, 2 were tonic-off, and 2 were unclassified. Recordings from tonic-on fibers were obtained near tonic-on pretectal cells, typically in the most dorsal light-responsiveness region of the pretectum. These fibers were activated by single pulse electrical stimulation of the optic chiasm. The mean receptive field center diameter of 6 tonic-on fibers was 10.1°, or about a factor of 3 less than that of pretectal tonic-on cells. The mean conduction velocity of 14 tonic-on fibers was 3.1 m/s.We argue that the tonic-on cells of the PO serve to integrate signals from tonic-on center retinal ganglion cells with adjacent receptive fields to provide signals for constriction of the pupil to neurons in the oculomotor nucleus.     

Ultra-slow oscillatory neuronal activity in the rat olivary pretectal nucleus: comparison with oscillations within the intergeniculate leaflet

Oscillations with a period between 1 and 4 min have been previously observed in many visual system structures. To determine whether similar oscillations in neuronal firing also exist in rat pretectum, recordings of neuronal activity were made with standard extracellular recording methods in vivo . Oscillations with a mean period of ∼140 s were identified in 127 recording sites in olivary pretectal nucleus (OPT). Prolonged iontophoretical current ejection of bicuculline, an antagonist of GABA_A receptors, increased the firing rate but did not disrupt the oscillatory pattern of activity. This suggests that rhythmic activity of OPT neurons is either intrinsic to the nucleus or driven by rhythmic excitatory input. It is worth noting that oscillations within OPT were synchronized with the above-described oscillatory activity in the ipsilateral intergeniculate leaflet (IGL). In the case of simultaneous double recordings from OPT and contralateral OPT or IGL, oscillations were uncorrelated. Our findings suggest functional coupling of the OPT with ipsilateral IGL, and imply that OPT, besides its well established role in pupil constriction, might be involved in modulation of the neuronal mechanism of the circadian timing system, as was suggested previously. Alternatively, IGL might be involved in pupil diameter regulation.     

The pretectal olivary nucleus of the cat: evidence for a two-tailed structure

The pretectal olivary nucleus of the cat was reconstructed from Nissl and myelin stained tissue. The olivary nucleus is found to have a medially placed head region and two tail regions which extend obliquely from the head region. The retino-olivary projection was also analyzed using the anterograde autoradiographic tracing method. Retinal input is found bilaterally over both the head of the nucleus and the tail regions. The distribution of silver grains, when compared bilaterally, appears slightly more dense over the contralateral nucleus.     

Comparison of the distributions of urocortin-containing and cholinergic neurons in the perioculomotor midbrain of the cat and macaque

Urocortin is a novel neurotransmitter that appears to play a role in eating and drinking behavior. Most urocortin-positive (urocortin(+)) neurons in rodents are found in the cytoarchitecturally defined Edinger-Westphal nucleus (EW). However, the EW is traditionally described as the source of the preganglionic parasympathetic outflow to the ciliary ganglion. We examined the distribution of urocortin(+) cells and motoneurons by use of immunohistochemical staining for this peptide and for choline acetyltransferase (ChAT) in macaque monkeys, in which most preganglionic motoneurons inhabit the EW, and in cats, in which most do not. In both species, lack of overt double labeling indicated that the ChAT(+) and urocortin(+) cells are separate populations. In the monkey, most nonoculomotor ChAT(+) neurons were found within the EW. In contrast, urocortin(+) cells were distributed mainly between the oculomotor nuclei and in the supraoculomotor area. In the cat, most nonoculomotor ChAT(+) cells were located in the supraoculomotor area and anteromedian nucleus. Few were present in the cat EW. Instead, this nucleus was filled with urocortin(+) cells. These results highlight the fact the term EW has come to indicate different nuclei in different species. Consequently, we have adopted the identifiers preganglionic (EW(PG)) and urocortin-containing (EW(U)) to designate the cytoarchitecturally defined EW nuclei in monkeys and cats, respectively. Furthermore, we propose a new open-ended nomenclature for the perioculomotor (pIII) cells groups that have distinctive projections and neurochemical signatures. This will allow more effective scientific discourse on the connections and function of groups such as the periculomotor urocortin (pIII(U)) and preganglionic (pIII(PG)) populations.     

The urocortin 1 neurocircuit: ethanol-sensitivity and potential involvement in alcohol consumption

One of the hallmarks of alcoholism is continued excessive consumption of alcohol-containing beverages despite the negative consequences of such behavior. The neurocircuitry regulating alcohol consumption is not well understood. Recent studies have shown that the neuropeptide urocortin 1 (Ucn1), a member of the corticotropin-releasing factor (CRF) family of peptides, could be an important player in the regulation of alcohol consumption. This evidence is accumulated along three directions of research: (1) Ucn 1-containing neurons are extremely sensitive to alcohol; (2) the Ucn1 neurocircuit may contribute to the genetic predisposition to high alcohol intake in mice and rats; (3) manipulation of the Ucn1 system alters alcohol consumption and sensitivity. This paper reviews the current knowledge of the Ucn1 neurocircuit and the evidence for its involvement in alcohol-related behaviors, and proposes a mechanism for its involvement in the regulation of alcohol consumption.     

Premotor circuits controlling eyelid movements in conjunction with vertical saccades in the cat: I. The rostral interstitial nucleus of the medial longitudinal fasciculus

Saccadic eye movements in the vertical plane are controlled by the rostral interstitial nucleus of the medial longitudinal fasciculus (riMLF) and the interstitial nucleus of Cajal. Eye movements in the vertical direction are accompanied by concurrent upper eye lid movements. These gaze-related lid movements are produced by the levator palpebrae superioris muscle, whose motoneurons are located in the caudal central subdivision (CCS) of the oculomotor nucleus. The neural circuits that direct such gaze-related lid movements were examined by use of both conventional and dual neuronal tracing methods in the cat. Injections of wheat germ agglutinin-conjugated horseradish peroxidase (WGA-HRP) into the area of the CCS revealed a distinctive subset of retrogradely labeled neurons located in the caudomedial portion of the riMLF. This subset of riMLF neurons was not labeled when injections were localized within the oculomotor nucleus proper, without involving the CCS. Injections of biotinylated dextran amine (BDA) that included this caudomedial riMLF region anterogradely labeled axons that projected profusely throughout the CCS. Labeled terminals were seen in close association with retrogradely labeled levator palpebrae motoneurons, which were primarily found contralateral to WGA-HRP muscle injections. Ultrastructural examination revealed that most BDA-labeled terminals contained clear spherical vesicles and formed asymmetrical synaptic contacts, primarily on the proximal dendrites of WGA-HRP-labeled motoneurons. A few had pleiomorphic vesicles. In summary, these results strongly suggest that the caudomedial part of the cat riMLF is a premotor center that monosynaptically controls lid movements in conjunction with vertical saccades.     

Vestibular nucleus projections to the Edinger-Westphal and anteromedian nuclei of rabbits

The Edinger-Westphal nucleus (EW), anteromedian nucleus (AM) and adjacent neurons in the ventral tegmental area (VTA) are sources of preganglionic parasympathetic innervation of intraocular smooth muscle, including blood vessels, pupillary muscle and the ciliary body in mammals. They also have central connections that are believed to affect parasympathetic outflow indirectly. This study utilized anterograde transport of biotinylated dextran amine and Phaseolus vulgaris leucoagglutinin to demonstrate direct projections from the vestibular nuclei to the Edinger-Westphal and anteromedian nuclei in rabbits. The rabbit AM and adjacent VTA contain moderate to intensely choline acetyltransferase (ChAT)-immunopositive neurons. The rabbit EW, by contrast, is nearly devoid of ChAT-immunopositive neurons. Vestibular nucleus projections to these regions originate from all levels of the superior, medial and lateral vestibular nuclei, but do show topographic organization. The densest terminations were observed in AM and the ventral and central aspects of EW. The projections to AM terminate in both ipsilaterally and in a narrow paramedian region. Predominantly ipsilateral terminations were observed in VTA. Terminations on ChAT-positive cells in AM and VTA were verified in three rabbits. It is suggested that projections to some intensely ChAT-positive AM and VTA neurons may be a substrate for vestibular influences on lens accommodation, pupillary constriction and regulation of intraocular circulation during changes in posture and gravitoinertial challenges. The projections to ChAT-negative (and weakly immunoreactive) cells in AM, VTA and EW, on the other hand, are likely to contribute vestibular signals to a variety of motor responses via descending pathways.     

Direct projections from the anterior pretectal nucleus to the dorsal accessory olive in the cat: an anterograde and retrograde WGA-HRP study

Direct projections from the anterior pretectal nucleus (APN) to the dorsal accessory olive (DAO) were found in the cat by the anterograde and retrograde WGA-HRP methods. The dorsal or the ventral portions of the rostral half of the APN pars compacta send fibers respectively to the lateral or the medial portions of the whole rostrocaudal extent of the DAO. These APN-DAO fibers can be considered to play roles in some somatomotor mechanisms.     

Nitrous oxide-induced c-Fos expression in the rat brain

Induction of c-Fos has previously been used to map locations of cells in the central nervous system (CNS) that are activated by ethanol administration. Only a few studies examining a restricted range of CNS areas have identified brain areas activated by nitrous oxide (N(2)O). Because ethanol and N(2)O have overlapping physiological, psychological and behavioral effects, we hypothesized that these drugs act on similar sites in the CNS. To test this hypothesis, we assessed c-Fos-like immunoreactivity in brain slices from male Long-Evans rats that received a 2-h exposure of 0, 20, 40 or 60% N(2)O (n=5 each) immediately prior to sacrifice. N(2)O administration produced significant (P<0.05) dose-related increases of c-Fos expression in several forebrain regions, including the hypothalamic supraoptic and paraventricular nuclei, the thalamic paraventricular nucleus, the amygdala, and in retrosplenial cortex. In the midbrain, N(2)O caused significant dose-related c-Fos expression in the Edinger-Westphal nucleus. Finally, the pontine locus coeruleus, and two medullary regions, the nucleus of the solitary tract and ventrolateral medulla, also showed significant dose-related N(2)O-induced c-Fos expression. Most of the brain areas identified as targets of N(2)O are also activated by ethanol administration. The overlapping pattern of c-Fos induced by ethanol and N(2)O suggests that these drugs may cause comparable central activity by acting on similar neuronal pathways.     

Premotor circuits controlling eyelid movements in conjunction with vertical saccades in the cat: II. interstitial nucleus of Cajal

Vertical saccadic eye movements are accompanied by concurrent eyelid movements in the same direction. The interstitial nucleus of Cajal (InC) controls eye position for vertical eye movements and may also control saccade-related lid position as well. This study investigates whether the InC serves as a premotor center for eyelid saccades, by employing dual-tracer methods in cats to label both the projections of the InC and the motoneurons supplying the levator palpebrae superioris (LPS) muscle, which lie in the caudal central subdivision (CCS) of the oculomotor complex. Injections of biotinylated dextran amine (BDA) into the InC anterogradely labeled axons that terminated bilaterally throughout the CCS and in the oculomotor nuclei proper. Labeled terminals lay in close association with labeled LPS motoneurons, which were retrogradely labeled following injections of wheat germ agglutinin-conjugated horseradish peroxidase (WGA-HRP) into the muscle. Ultrastructural investigation revealed that most terminals contained spherical vesicles and formed asymmetric synaptic contacts with the labeled motoneurons. These results strongly suggest that the InC monosynaptically controls lid movements in conjunction with vertical eye movements, including saccades. To identify the neurons of origin for this pathway, WGA-HRP injections were centered in the CCS. These experiments indicate that lid and eye motoneurons may share a common source of bilateral InC input. Thus, a common vertical position signal may be employed to maintain the lid and eye at appropriate elevations during fixation, such that the lid sits just above the pupil, allowing unobstructed vision, but at the ready to protect the cornea.     

The cause of increased pupillary light reflex latencies in diabetic patients: the relationship between pupillary light reflex and visual evoked potential latencies.

In 42 diabetic patients the relationship between the latency of the pupillary light reflex and the pattern reversal visual evoked potential (P100) was examined. Fifty-five percent of diabetic patients had pupillary light reflex latencies above the normal range. In 19% the visual evoked potentials were prolonged when compared to the normal range. Latencies of pupillary light reflexes and VEPs showed no correlation. There was a minimal correlation between the presence of retinopathy and prolongation of both the pupillary light reflex and the visual evoked response latency (kappa coefficients respectively: 0.31, P less than 0.01 and 0.36, P less than 0.02). The presence of an increased pupillary light reflex latency was positively correlated with a reduced respiratory sinus arrhythmia (kappa coefficient: 0.58, P less than 0.0001). Increased VEP latencies showed no correlation with signs of cardiovascular autonomic neuropathy. We conclude that the afferent optic pathway can be affected in diabetic patients. However, prolongation of pupillary light reflex latency in diabetic patients is primarily due to an efferent pupillary defect and represents parasympathetic dysfunction.     

The nucleus pretectalis principalis: A pretectal structure hidden in the mammalian thalamus

A defining feature of the amniote tecto-fugal visual pathway is a massive bilateral projection to the thalamus originating from a distinct neuronal population, tectal ganglion cells (TGCs), of the optic tectum/superior colliculus (TeO/SC). In sauropsids, the thalamic target of the tecto-fugal pathway is the nucleus rotundus thalami (Rt). TGCs axons collateralize en route to Rt to target the nucleus pretectalis principalis (PT), which in turn gives rise to bilateral projection to the TeO. In rodents, the thalamic target of these TGCs afferents is the caudal division of the pulvinar complex (PulC). No pretectal structures in receipt of TGC collaterals have been described in this group. However, Baldwin et al. (Journal of Comparative Neurology, 2011;519(6):1071–1094) reported in the squirrel a feedback projection from the PulC to the SC. Pulvino-tectal (Pul-T) cells lie at the caudal pole of the PulC, intermingled with the axonal terminals of TGCs. Here, by performing a combination of neuronal tracing, immunohistochemistry, immunofluorescence, and in situ hybridization, we characterized the pattern of projections, neurochemical profile, and genoarchitecture of Pul-T cells in the diurnal Chilean rodent Octodon degus. We found that Pul-T neurons exhibit pretectal, but not thalamic, genoarchitectonical markers, as well as hodological and neurochemical properties that match specifically those of the avian nucleus PT. Thus, we propose that Pul-T cells constitute a pretectal cell population hidden within the dorsal thalamus of mammals. Our results solve the oddity entailed by the apparent existence of a noncanonic descending sensory thalamic projection and further stress the conservative character of the tectofugal pathway.     

Spatial distribution of evoked potentials in the inferior olivary nucleus by stimulation of the visual afferents in the rat

Visual pathways (optic disc, optic nerve and pretectal regions) were electrically stimulated and evoked potentials were explored throughout the inferior olive in the anesthetized rat. Responsive areas were identified as the caudal half of the dorsal cap, nucleus beta and the most caudal region of subnucleus c of the medial accessory olive. No field potentials were identified in the rostral half of the dorsal cap, its ventrolateral outgrowth or the dorsomedial cell column. Contralateral retinal afferents were only effective all over the responsive areas.     

Horizontal optokinetic nystagmus in unilaterally enucleated pigmented rats: role of the pretectal commissural fibers.

Monocular enucleation reduces the asymmetry of horizontal optokinetic nystagmus (H-OKN) in afoveate mammals by increasing responses to naso-temporal visual stimulation. The origin of these larger responses was investigated in adult pigmented rats monocularly enucleated as neonates or as adults by analyzing retinal and commissural projections to the deafferented nucleus of the optic tract (NOT) and the functional role of this nucleus before and after section of the posterior commissure. Anatomically, monocular enucleation reduces the volume of the contralateral deafferented NOT. Anterograde tracers injected in the intact eye reveal a crossed projection of the retina to the NOT and to the dorsal (DTN) and medial (MTN) terminal nuclei of the accessory optic system as in normal rats. In addition, there is an uncrossed projection to the MTN in the rats enucleated as neonates. Retrograde tracer injected in the deafferented NOT confirms the absence of an uncrossed retinal projection but reveals connections between both NOT via the posterior commissure as in normal rats. Electrophysiologically, the larger naso-temporal optokinetic responses in monocularly enucleated rats return to normal after posterior commissurotomy. This study demonstrates that no anatomical remodelling takes place to increase naso-temporal responses in monocularly enucleated rats. The larger responses must then result from functional changes. The role of exclusive contralateral projections of the retina to the NOT and of the commissural connections in mediating the asymmetry of the optokinetic nystagmus in afoveate mammals is discussed.     

Laminar distribution of tectal, parabigeminal and pretectal inputs to the primate dorsal lateral geniculate nucleus: Connectional studies in Galago crassicaudatus

We have studied the distribution of 3 extraretinal, subcortical inputs to the dorsal lateral geniculate nucleus of the prosimian primate Galago. Our connectional findings reveal that axons arising from the superior colliculus and the parabigeminal nucleus influence the W-cell system via their innervation of the two small-celled geniculate laminae (internal and external koniocellular) and the interlaminar zones; parabigeminal axons also innervate each of the 4 non-tectally innervated layers. Pretectal axons, on the other hand, distribute mainly to the parvocellular laminae and thus influence primarily the X-cell system.     

The pretectal complex in the opossum: projections from the striate cortex and correlation with retinal terminal fields

Two terminal fields were revealed in the pretectal complex of the opossum by the Fink-Heimer method after striate cortical lesions. A rostral field is located within a rostrolateral strip of the compact part of the anterior pretectal nucleus, where a partial topographic arrangement of this projection is present. A caudal field is located within the sub-brachial nucleus of the optic tract, located between the brachium of the superior colliculus and the posterior pretectal nucleus. The corticotopic projection to this field is mirror-symmetric to that found in the superior colliculus and overlaps a bilateral projection from the retina. Based on hodological evidence, it is concluded that the nucleus of the optic tract in the opossum can be subdivided in (a) an intrabrachial nucleus receiving a direct projection from the contralateral retina and (b) a sub-brachial nucleus receiving projections from both retinae and from the striate cortex. The pretectal complex, as the superior colliculus, can be anatomically subdivided in a superficial region receiving visual input (theoptic pretectum) and a deep region only remotely connected to the visual system. The optic pretectum, however, differs from the superior colliculus in displaying a multiple-map arrangement within its constituent nuclei, instead of a single continuous representation of the visual field.