CR-positive Neurons Research Articles

Ontogeny of calcium-binding proteins in the cingulate cortex of the guinea pig: The same onset but different developmental patterns

This paper compared the density of calbindin D28k (CB), calretinin (CR) and parvalbumin (PV) containing neurons in prenatal, newborn and postnatal periods in the cingulate cortex (CC) of the guinea pig as an animal model. The distribution and co-distribution among calcium-binding proteins (CaBPs) was also investigated during the entire ontogeny. The study found that CB-positive neurons exhibited the highest density in the developing CC. The CC development in the prenatal period took place with a high level of CB and CR immunoreactivity and both of these proteins reached peak density during fetal life. The density of PV-positive neurons, in contrast to CB and CR-positive neurons, reached high levels postnatally. The observed changes of the CaBPs-positive neuron density in the developing CC coincide with developmental events in the guinea pig. E.g. the eyes opening moment may be preceded by elevated levels of CB and CR at E50, whereas high immunoreactivity of PV from P10 to P40 with a peak at P20 may indicate the participation of PV in enhancement of the inhibitory cortical pathway maturation.

A Subset of Palisade Endings Only in the Medial and Inferior Rectus Muscle in Monkey Contain Calretinin.

PurposeTo further chemically characterize palisade endings in extraocular muscles in rhesus monkeys.MethodsExtraocular muscles of three rhesus monkeys were studied for expression of the calcium-binding protein calretinin (CR) in palisade endings and multiple endings. The complete innervation was visualized with antibodies against the synaptosomal-associated protein of 25 kDa and combined with immunofluorescence for CR. Six rhesus monkeys received tracer injections of choleratoxin subunit B or wheat germ agglutinin into either the belly or distal myotendinous junction of the medial or inferior rectus muscle to allow retrograde tracing in the C-group of the oculomotor nucleus. Double-immunofluorescence methods were used to study the CR content in retrogradely labeled neurons in the C-group.ResultsA subgroup of palisade and multiple endings was found to express CR, only in the medial and inferior rectus muscle. In contrast, the en plaque endings lacked CR. Accordingly, within the tracer-labeled neurons of the C-group, a subgroup expressed CR.ConclusionsThe study indicates that two different neuron populations targeting nontwitch muscle fibers are present within the C-group for inferior rectus and medial rectus, respectively, one expressing CR, one lacking CR. It is possible that the CR-negative neurons represent the basic population for all extraocular muscles, whereas the CR-positive neurons giving rise to CR-positive palisade endings represent a specialized, perhaps more excitable type of nerve ending in the medial and inferior rectus muscles, being more active in vergence. The malfunction of this CR-positive population of neurons that target nontwitch muscle fibers could play a significant role in strabismus.

Calretinin-immunoreactive neurons in the claustrum of the guinea pig

ABSTRACTBackground: The claustrum is present in all mammalian species. Many aspects of its morphology and function remain subject to debate. It has been suggested that calcium-binding proteins influence neuronal activity in many structures.Objective: The aim was to examine CR immunoreactivity in the claustrum of guinea pigs and compare it with that in other mammals.Design: CR immunoreactivity in the claustrum was analyzed in five guinea pigs using immunohistochemical techniques. The distribution of CR-positive neurons in the dorsal claustrum and endopiriform nucleus (END), and their morphological characteristics were described.Results: CR immunostaining in the dorsal claustrum and END of the guinea pig consisted of CR-positive neurons, fibers, and puncta of varying labeling density. The majority of these neurons displayed small- or medium-sized round, oval, and multipolar cell bodies with aspiny dendrites. CR-ir neurons prevailed in the periphery of both divisions. The dorsal claustrum was seen to have a weakly CR-ir central core surrounded by a rim of moderately positive neuropil.Conclusion: CR immunoreactivity in the guinea pig claustrum was not as diffuse as in higher mammals, but non-uniformity was evident. CR-ir neurons are distributed in both the dorsal claustrum and END, mostly in the periphery.

Calretinin as a Marker for Premotor Neurons Involved in Upgaze in Human Brainstem

Eye movements are generated by different premotor pathways. Damage to them can cause specific deficits of eye movements, such as saccades. For correlative clinico-anatomical post-mortem studies of cases with eye movement disorders it is essential to identify the functional cell groups of the oculomotor system in the human brain by marker proteins. Based on monkey studies, the premotor neurons of the saccadic system can be identified by the histochemical markers parvalbumin (PAV) and perineuronal nets in humans. These areas involve the interstitial nucleus of Cajal (INC) and the rostral interstitial nucleus of the medial longitudinal fascicle (RIMLF), which both contain premotor neurons for upgaze and downgaze. Recent monkey and human studies revealed a selective excitatory calretinin (CR)-positive input to the motoneurons mediating upgaze, but not to those for downgaze. Three premotor regions were identified as sources of CR input in monkey: y-group, INC and RIMLF. These findings suggest that the expression pattern of parvalbumin and CR may help to identify premotor neurons involved in up- or downgaze. In a post-mortem study of five human cases without neurological diseases we investigated the y-group, INC and RIMLF for the presence of parvalbumin and CR positive neurons including their co-expression. Adjacent thin paraffin sections were stained for the aggrecan (ACAN) component of perineuronal nets, parvalbumin or CR and glutamate decarboxylase. The comparative analysis of scanned thin sections of INC and RIMLF revealed medium-sized parvalbumin positive neurons with and without CR coexpression, which were intermingled. The parvalbumin/CR positive neurons in both nuclei are considered as excitatory premotor upgaze neurons. Accordingly, the parvalbumin-positive neurons lacking CR are considered as premotor downgaze neurons in RIMLF, but may in addition include inhibitory premotor upgaze neurons in the INC as indicated by co-expression of glutamate decarboxylase in a subpopulation. CR-positive neurons ensheathed by perineuronal nets in the human y-group are considered as the homolog premotor neurons described in monkey, projecting to superior rectus (SR) and inferior oblique (IO) motoneurons. In conclusion, combined immunostaining for parvalbumin, perineuronal nets and CR may well be suited for the specific identification and subsequent analysis of premotor upgaze pathways in clinical cases of isolated up- or downgaze deficits.

Diverse interneuron populations have highly specific interconnectivity in the rat piriform cortex

Previous studies have suggested that the patterns of innervation and high interconnectivity of the piriform cortex (PC) provide for strong olfactory hippocampal memory; however, these same attributes may create high seizurogenic tendencies. Thus, understanding this wiring is important from a physiological and pathophysiological perspective. Distinct interneurons expressing differing calcium binding proteins (CBPs), parvalbumin (PV), calbindin (CB), and calretinin (CR), have been shown to exist in PC. However, a comprehensive examination of the distribution and innervation patterns of these neurons has not been done. Thus the purpose of this study was to combine the analysis of the CBP cell localization with analysis of their innervation patterns. Each type was differentially localized in the three layers of the PC. Only CR-positive neurons were found in layer 1. PV and CB are coexpressed in layers 2-3, most expressing both PV and CB. A morphological estimate of the dendritic extent for each subtype showed that PV and PV/CB cells demonstrated equally wide, horizontal and vertical arborizations, whereas CB cells had wide horizontal and restricted vertical arborizations. CR cells had restricted horizontal and very long vertical arborizations. Postsynaptic morphological targeting was also found to be specific, namely, PV(+) and PV/CB(+) nerve terminals (NTs) innervate perisomatic regions of principal cells. CR(+) NTs innervate only dendrites of principal cells, and CB(+) NTs innervate both somata and dendrites of principal cells. These data show highly complex innervation patterns for all of the CBP interneurons of the PC and form a basis for further studies in the plasticity of this region.

Neuroprotective activity of omega-3 fatty acids against epilepsy-induced hippocampal damage: Quantification with immunohistochemical for calcium–binding proteins

To investigate whether n-3 polyunsaturated fatty acids ( n-3 PUFAs) would promote different morphological changes in the hippocampal formation of rats with epilepsy, we performed an immunocytochemical study using parvalbumin (PV) and calretinin (CR) distribution as a marker. Animals subjected to the experimental model of epilepsy with a single dose of pilocarpine were randomly divided into the following groups: animals with epilepsy treated daily with vehicle (EV) and animals with epilepsy treated daily with 85 mg/kg n-3 PUFAs (EW). Control animals administered saline were also randomly divided into two other groups: animals treated daily with vehicle (CV) and animals treated daily with 85 mg/kg n-3 PUFAs (CW). A larger number of PV-positive neurons were observed in EW when compared with EV, CV, and CW. Similarly, there were significantly more CR-positive neurons in EW than in EV. These findings demonstrate that omega-3 fatty acids prevent status epilepticus-associated neuropathological changes in the hippocampal formation of rats with epilepsy.

Decreased expression of calretinin in the cerebral cortex and hippocampus of SOD1 G93A transgenic mice

In the present study, we investigated the changes of calretinin (CR) expression in the central nervous system of SOD1 G93A transgenic mice as an in vivo model of amyotrophic lateral sclerosis (ALS). In wild-type SOD1 (wtSOD1) transgenic mice, many CR-immunoreactive neurons were found in all cortical regions. In the cerebral cortex of SOD1 G93A transgenic mice, the number and staining intensity of CR-positive neurons were decreased. In the hippocampal formation, layer-specific alterations in the staining intensity of CR-immunoreactive neurons were observed in the CA1–3 areas and dentate gyrus. In wtSOD1 transgenic mice, CR-immunoreactive neurons with long processes were found in the stratum oriens and stratum radiatum of CA1–3 areas, and heavily stained band-like molecular layer was prominent in the dentate gyrus. CR immunoreactivity was decreased in each layer of CA1–3 areas and dentate gyrus of SOD1 G93A transgenic mice. The first demonstration of decreased immunoreactivity for CR in the cerebral cortex and hippocampus of SOD1 G93A transgenic mice may provide insights into the pathogenesis of motor neuron degeneration in human ALS although further quantitative studies are needed.

Differential effects of aging on the distribution of calcium-binding proteins in a pretectal nucleus of the chicken brain

The nucleus pretectalis (PT) of birds is an ovoid-shaped visuomotor cell group of the pretectum that receives tectal input and projects back to the optic tectum. We performed immunohistochemical single- and double-labeling to determine the distribution and abundance of neurons containing three calcium-binding proteins, parvalbumin (PV), calretinin (CR), and calbindin (CB), in the PT in chickens at three ages. We found that PV-positive and CR-positive cells co-localize and are largely found in the outer part of PT at all ages. The GluR4 subunit of the AMPA-type glutamate receptor was selectively localized to these neurons. CB-positive neurons, however, were largely absent from the PT in young and adult chickens. The abundance of PV-positive and CR-positive neurons in PT in old birds was indistinguishable from that in the younger birds, but CB-positive perikarya were 10–20-fold more common than in young birds, and were again mainly found in the outer part of PT. The overall abundance of neurons in PT was reduced to about 50% of its former abundance in the old birds, with this loss restricted to the central part of the nucleus. These data indicate that a cell loss process develops in PT as birds age, that parvalbuminergic and calretinergic neurons resist this process, and that this process is associated with increased expression of CB.

NADPH-diaphorase and calcium binding proteins in the trigeminal nucleus oralis of rats

We have examined the distribution of nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) and the calcium binding proteins (CBPs), calbindin D-28k (CB), calretinin (CR) and parvalbumin (PV), in the trigeminal nucleus oralis (Sp5O). NADPH-d was detected by histochemistry while CBP was detected by immunohistochemistry. NADPH-d-positive neurons were distributed in the medial rostro-dorsomedial part (RDMsp5O) and dorsomedial part (DMsp5O) of Sp5O, and the rostrolateral part of the nucleus of the solitary tract (NTS). CB- and CR-positive neurons were mainly distributed in the dorsal part of Sp5O. In contrast, PV-positive neurons were mainly distributed in the ventral part of Sp5O. NADPH-d colocalized with CB (40%) and CR (20%) but not with PV in neurons of DMsp5O/NTS. The mean cell sizes of neurons in RDMsp5O were larger than those in DMsp5O/NTS. PV-positive neurons were larger than NADPH-d-positive neurons. NADPH-d-, CB- and CR-positive neurons were generally small in RDMsp5O and DMsp5O/NTS. Few neurons were retrogradely labeled in RDMsp5O and DMsp5O from the thalamus, when numerous labeled neurons were in the principal and interpolar nuclei. These data indicate that NADPH-d histochemistry and CB, CR and PV immunohistochemistry identify a discrete cell population in Sp5O. Those labeled neurons in RDMsp5O and DMsp5O/NTS were considered to be involved in sensorimotor reflexive function of the intra-oral structures.

Ruffini endings are absent from the periodontal ligament of trkB knockout mice

To clarify the role of neurotrophin receptors in the development of Ruffini endings, periodontal ligaments and trigeminal ganglia of trkA, trkB, and trkC knockout mice were immunostained for protein gene product 9.5 (PGP 9.5), calcitonin gene-related peptide (CGRP), parvalbumin (PV), and calretinin (CR). Innervation patterns of PGP 9.5- and CGRP-immunoreactive fibers were examined in the periodontal ligament of the knockout mice. PGP 9.5-positive fibers in the incisal periodontal ligaments of trkA and trkC knockout mice form Ruffini endings distinguished by dendritic ramifications and branches. However, Ruffini endings were not present in the periodontal ligament of trkB knockout mice. Only free nerve endings were observed in tissue of trkB knockout mice. Compared with trkA and trkC knockouts, the proportion of CR-positive neurons in mandibular and maxillary regions of the trigeminal ganglion of trkB knockout mice is decreased. These findings indicate that the development of periodontal Ruffini endings is regulated by trkB-dependent and CR-coexpressing neurons.

Calbindin-D28k and calretinin immunoreactive neurons in the olfactory bulb of the musk shrew, Suncus murinus

The distribution, morphological features, and postnatal development of calbindin-D28k (CB) and calretinin (CR) immunoreactive neurons in the main olfactory bulb (MOB) of the musk shrew, Suncus murinus, were studied by immunostaining to determine the degree of colocalization of CB and CR, and the relationship of CB and CR to neuron development in the MOB of animals of the order Insectivora. In adults, CB-positive neurons were identified as periglomerular and perinidal cells in the periglomerular region, as superficial short-axon cells in the external plexiform layer, and as four types of interneurons (Cajal, horizontal, Golgi, and bitufted cells) in the mitral cell, internal plexiform, and granule cell layers. CR-positive neurons were identified as projection neurons (tufted and mitral cells) and interneurons (periglomerular, perinidal, and granule cells). On postnatal days 1 and 3, CB-positive neurons revealed numerous processes finely arborized near the somata, and were morphologically unidentifiable. At the same time, CR-positive neurons were identified as young periglomerular and granule cells, and as migrating bipolar cells extending leading processes with growth cones in each layer of the MOB and the subependymal layer between the anterior lateral ventricle and the center of the MOB. On postnatal day 28, mature CB-positive and CR-positive interneurons were distributed in their corresponding layers, whereas migrating CR-positive bipolar cells were rarely detected. No cells colocalized CB and CR. The results suggest that perinidal cells in the shrew MOB may develop postnatally, together with glomerular and granule cells. We suggest that CB is associated with mechanisms of the outgrowth of neuronal processes, whereas CR is involved in mechanisms of cell migration and outgrowth of neuronal processes, in some types of neurons in the developing stage of the shrew MOB.

Relative sparing of calretinin containing neurons in the substantia nigra of 6-OHDA treated rat Parkinsonian model

A certain calcium binding protein (CaBP) has been known to exert a neuroprotective effect in various neurodegenerative diseases. Using the 6-OHDA induced rat Parkinsonian model, we examined if calretinin (CR), one of CaBP family, could play the similar role in the Parkinson's disease because CR is profusely localized in dopaminergic neurons of the substantia nigra pars compacta (SNPC) of the rat. Employing immunohistochemical analyses, we found that the survival rate of CR neurons was significantly higher than that of tyrosine hydroxylase (TH) neurons in the SNPC of the Parkinsonian rat. Furthermore double-labeled fluorescent microscopy revealed that almost all surviving TH neurons were also positive to CR. Our data suggest that CR-positive neurons are less vulnerable to 6-OHDA and CR in the dopaminergic neurons may have a protective function for survival of these neurons in the experimentally induced Parkinsonian rat.

Differential effect of thyroid hormone deficiency on the growth of calretinin-expressing neurons in rat spinal cord and dorsal root ganglia.

The development of spinal cord or dorsal root ganglia neurons expressing calretinin (CR) was studied in thyroid hormone-deficient rats. Immunocytochemical and morphometric analyses showed that the hypothyroidism induced a significant decrease in the number and size of immunoreactive neurons in the spinal cord, as well as stunted growth and arborization of the axons and dendrites. These alterations were observed at different embryonic ages and persisted during the whole postnatal life. In adult hypothyroid rats, the mean number of CR-positive neurons per spinal cord section (31.2 +/- 2.3 in laminae I and II and 30.5 +/- 5.5 in laminae III-X) was significantly decreased (P < 0.001 and P = 0.024, respectively) compared with adult normal rats (68.7 +/- 8.9 and 50.0 +/- 11.0, respectively). In the peripheral nervous system, hypothyroidism altered the growth of sensory neurons expressing CR protein mainly during embryonic life. In comparison with normal rats, hypothyroid embryonic animals showed not only reduced cell size but also a significantly decreased percentage of CR-positive neurons (6.6 +/- 0. 9% in normal, 2.1 +/- 0.3% in hypothyroid rats, P < 0.001). In contrast, although the size of neurons was reduced in hypothyroid young and adult rats, there was no reduction in the percentage of CR-positive neurons. These results showed that thyroid hormone deficiency altered differentially the development of neurons expressing CR protein in the central and peripheral nervous systems. This suggests that central and peripheral neurons are heterogeneous in their sensitivity to thyroid hormone.

Postnatal development of calretinin- and parvalbumin-positive interneurons in the rat neostriatum: an immunohistochemical study.

On the basis of cytochemical and morphologic differences, two classes of gamma-aminobutyric acidergic (GABAergic) interneurons expressing calcium-binding proteins have been identified in the striatum of adult animals: neurons expressing either parvalbumin (PV) or calretinin (CR). The function of these calcium-binding proteins is not clear, however, they are associated with distinct classes of inhibitory interneurons within the adult neostriatum. By using immunocytochemical techniques, we analyzed the postnatal maturation and the spatiotemporal distribution of PV- and CR-positive neurons in the rat neostriatum compared with a third class of interneurons characterized by the expression of the acetylcholine-synthesizing enzyme, choline acetyltransferase (ChAT). PV-positive cells appeared initially on postnatal day 9 in the lateral region of the striatum. During postnatal weeks 2 and 3, the numbers of PV-positive neurons increased, and this cell population spread progressively in a lateromedial direction. In contrast, CR-expressing neurons were present at birth. During the first few days after birth, the number of CR-immunoreactive cells increased, reaching a peak on postnatal day 5 before declining during the following 2 weeks. A mediolateral gradient was evident temporarily. ChAT-containing neurons were detectable at birth in the lateral striatum. During postnatal weeks 1 and 2, the neurons matured along a lateral-to-medial gradient. The results indicate that the maturation of striatal interneurons is regulated differentially during postnatal development, resulting in a distinct spatiotemporal genesis of phenotypes. The sequential expression of CR and PV suggests a stage-dependent development of subsets of inhibitory interneurons and, hence, the stage-dependent maturation of functionally distinct inhibitory circuits within the neostriatum.

Calretinin-immunoreactivity during postnatal development of the rat isocortex: a qualitative and quantitative study.

Postnatal development of the rat cortex is characterized by the gradual development of many calcium-dependent processes which demand a precise control of the intracellular levels of this cation; when the balance is disturbed, neuronal death ultimately ensues. Calretinin (CR), a calcium-binding protein, has been postulated to have neuroprotective capacity by buffering intracellular calcium. This putative relationship between CR and neuroprotection is still, however, a controversial issue. With a view to shedding further light on this subject, we studied the temporal and spatial distribution of CR in five different regions (the frontal- sensorimotor-, parietal-, temporal- and occipital region) of the rat cortex during postnatal development. Qualitative and quantitative immunocytochemistry of newborn, 5-, 10-, 15-, 20- and 30-day-old and adult rats revealed a profound increase in the density of the CR-positive neurons during the first two postnatal weeks in all regions examined. At the end of this period, CR-immunoreactive cells decreased sharply to adult levels. Cell classes exhibiting transient CR-immunoreactivity during the first two postnatal weeks included cells in layer I (amongst which were Cajal-Retzius cells), the subplate and pyramidal-like cells in the upper portion of layer V, most of them in the motor cortices. The above-described dynamics of CR expression were reflected also in the biochemical analysis performed (immunoblotting, ELISA). The temporal and spatial correlation with calcium-dependent events such as synaptogenesis, neurite elongation and remodelling in further support the contention that CR may play a neuroprotective role during postnatal development of the rat cortex.

Development of calretinin immunoreactivity in the neocortex of the rat.

The prenatal and postnatal development of calretinin (CR)-containing elements in the neocortex of the rat was analyzed using immunohistochemistry. CR immunoreactivity in the cortical anlage appeared early at embryonic day 14 (E14), with CR-positive neurons located in the primitive plexiform layer and in the emerging subplate and marginal zones. At later prenatal and early postnatal stages, these two layers showed the highest CR immunostaining in the cortex, and large numbers of cell bodies and fibers were immunostained. From postnatal day 3 (P3) onwards, CR immunostaining disappeared progressively from the subplate-layer VIb and the marginal zone-layer I, so that very few cells remained stained in these layers in the adult. In the cortical plate and prospective layers VIa to II-III, CR-positive neurons were seen at prenatal stages, their numbers increasing markedly during the first postnatal week. Most neurons showed undifferentiated nonpyramidal shapes, and matured during the second and third postnatal weeks, when the adult pattern of CR immunostaining was achieved. In addition, some pyramidal-like neurons in the infragranular layers and in layer II-III transiently expressed CR during the postnatal period, most notably between P3 and P12. Colocalization experiments performed at P0-P3 with antibodies against the neurotransmitter gamma-aminobutyric acid (GABA) showed that most nonpyramidal CR-positive neurons in the derivates of the cortical plate were also GABAergic during development. In contrast, large numbers of CR-containing neurons in the subplate and marginal zone were GABA-negative. The present results show that in addition to recording the early development of a subset of nonpyramidal neurons, CR is transiently expressed in certain GABA-negative populations of the subplate and marginal zone, and most likely in pyramidal neurons.

Loss of parvalbumin immunoreactivity defines selectively vulnerable thalamic reticular nucleus neurons following cardiac arrest in the rat.

The thalamic reticular nucleus (NRT) is one of the most vulnerable structures to selective neuronal damage both in human cardiac arrest patients and in experimental rodent global cerebral ischemia models. The detailed distribution of neuronal injury within the NRT was examined following 10-min cardiac arrest in the rat with conventional Nissl staining, 45Ca autoradiography and immunocytochemistry of the calcium binding proteins parvalbumin (PV) and calretinin (CR). While Nissl staining was almost unable to show the exact boundary of the nucleus and of the lesion, immunocytochemistry of PV proved to be the most useful index of the exact location and extent of neuronal loss in the NRT after ischemia. Calcium autoradiography was a sensitive method for detecting the lesion, and showed a similar distribution to the loss of PV staining, but did not give optimal spatial resolution. Quantitative analysis of PV staining at 7 days of recirculation demonstrated cell loss restricted to the lateral aspect of the middle segment of the NRT, identical with the distribution of large fusiform neurons in the somatosensory component of the nucleus. CR-positive neurons in the NRT were completely spared, although not all surviving neurons contained CR. These studies provide the first detailed characterization of the distribution of vulnerable neurons within the NRT after experimental ischemia and suggest that immunocytochemistry of PV is a useful tool for quantitative analysis of the lesion for use in further experiments to elucidate the mechanisms of selective vulnerability of the NRT.

Calretinin-immunoreactive neurons in the trigeminal and dorsal root ganglia of the rat

The cell body size of primary neurons were measured in the trigeminal (TG) and lumbar dorsal root ganglia (DRG) monochrome-stained for calretinin (CR)-like immunoreactivity. A trichrome stain for CR, carbonic anhydrase (CA) and tachykinin (TK) was also employed to estimate possible overlap of cellular distribution of these substances. In the DRG, the cell size spectrum of CR-positive cells was clearly bimodal; a greater proportion (84.1%) of CR-positive cells was distributed in the range ≧ 800 μm 2 with a mode between 1,500–1,600 μm 2, while a smaller proportion (14.8%) < 700 μm 2 with a mode of 400–500 μm 2. They were evenly distributed throughout the DRG. Although CR-positive TG neurons were smaller than similar DRG neurons, a bimodal distribution pattern remained unchanged. 94.6% of CR-positive cells measured 100–1,400 μm 2 with peak ranges of 200–300 μm 2 and 400–500 μm 2. Most of CR-positive cells in the ophthalmic division were 400 μm 2 or larger and small CR-positive cells ( < 400 μm 2 ) were concentrated in the maxillary and mandibular divisions. Most of CR-positive DRG cells showed CA activity (76.5%), while those with TK-immunoreactivity were rare (7.2%). In the TG, 38.4% of CR-positive cells were TK-positive. They were mostly smaller than 800 μm 2. On the other hand, CA was detected in 43.4% of CR-positive TG cells. Most of the TG cells co-expressing CR and CA were 400 μm 2 or larger. Simultaneous co-expression of TK and CA by the CR-positive cells was negligible. The cell bodies co-expressing CR and TK were more or less concentrated in the maxillary and mandibular divisions of the TG. The results suggest that CR-positive DRG cells are mostly proprioceptive, whereas similar TG cells include many small primary neurons innervating the visceral mucosa.

Calretinin-immunoreactive neurons in the primary visual cortex of dolphin and human brains

A new class of γ-aminobutyric acid (GABA)ergic neurons immmunoreactive to the calcium-binding protein calretinin (CR) was demonstrated in primary visual cortices of the bottlenose dolphin ( Tursiops truncatus) and humans ( Homo sapiens). Comparative analysis revealed several differences between dolphin and human visual cortex in the laminar distribution of CR-positive perikarya, although general typology of the immunoreactive CR-positive neurons was similar in both species. Thus, in both human and dolphin primary visual cortex almost all CR-positive neurons are non-pyramidal, either fusiform or bipolar cells, oriented with their long axis along the radial axis of the cortex. Large multipolar stellate cells were also observed in layers I and VI. The CR-positive neurons in the dolphin visual cortex are concentrated almost exclusively in layer I and, to a lesser extent, in layer II. In all other layers (IIIa,b, IIIc/ V and VI) of the dolphin visual cortex CR-positive neurons were only rarely seen. In the human primary visual cortex CR-positive neurons are located mainly in layers II, III and IVa, b, c, with considerably lower densities of these cells observed in layers V and VI. CR-positive neurons in layer I of the human visual cortex are represented by Cajal-Retzius horizontal cells, whereas no such cells were seen in layer I of the dolphin neocortex. The numerical density of CR-positive neurons in the dolphin primary visual cortex is significantly lower than in the same area of cortex in humans. However, percentages of total neuronal population by layers and overall in corrtex are significantly higher in the dolphin primary visual cortex (approximately 16.5%) in comparison with that in humans (approximately 0.9%).