Dietary plant oil supplemented with arachidonic acid and eicosapentaenoic acid affects the fatty acid composition and eicosanoid metabolism of Atlantic salmon (Salmo salar L.) during smoltification.

This study sought to investigate whether a "natural diet" (mimicking the fatty acid composition of freshwater aquatic insects eaten by salmon parr) during the freshwater (FW) life stage of pre-smolt Atlantic salmon (Salmo salar L.) affected red blood cells and gill fatty acid composition as well as eicosanoid metabolism in gill during smolting at different temperatures. Before being transferred to seawater (SW), salmon parr were fed with a modified (MO) diet containing vegetable oils (rapeseed, palm, and linseed oils) supplemented with eicosapentaenoic acid (EPA) and arachidonic acid (ARA) to completely replace the fish oil (FO). Fatty acid composition in red blood cells and gill tissues was determined before SW transfer and six weeks after. Additionally, the expression of genes associated with eicosanoid metabolism and Na+/K+-ATPase (NKA) activity in salmon gill was examined at different temperatures before SW transfer and 24 h after. The results showed the changes in fatty acid composition, including sum monounsaturated fatty acids (MUFAs), docosahexaenoic acid (DHA), ARA, EPA, and sum n-6 polyunsaturated fatty acids (n-6 PUFA) in both red blood cells and gill tissues at the FW stage were consistent with the fatty acid profiles of the supplied MO and FO fish diets; however sum EPA and DHA composition exhibited opposite trends to those of the FO diet. The proportion of ARA, EPA, and n-6 PUFA increased, whereas sum MUFAs and DHA decreased in the red blood cells and gill tissues of MO-fed fish compared to those fed with the FO diet at FW stage. Additionally, 5-lipoxygenase-activating protein (Flap) expression was downregulated in MO-fed fish prior to SW transfer. During the process of SW transfer at different temperatures, the MO diet remarkably suppressed NKAα1a expression in MO-fed fish both at 12 and 16 °C. The MO diet also upregulated phospholipase A2 group IV (PLA2g4) expression in gills at 8, 12, and 16 °C, but suppressed phospholipase A2 group VI (PLA2g6) expression in gills at 12 °C compared to FO-fed fish at 12 °C and MO-fed fish at 8 °C. The MO diet also upregulated Cyclooxygenase 2 (Cox-2) expression at 8 °C compared to FO-fed fish and increased Arachidonate 5-lipoxygenase (5-Lox) expression in MO-fed fish at 16 °C compared to both FO-fed fish at 16 °C and MO-fed fish at 8 °C. Our study also determined that both SW transfer water temperatures and diets during the FW period jointly influenced the mRNA expression of PLA2g4, PLA2g6, and Lpl, whereas 5-Lox was more sensitive to dietary changes. In conclusion, the MO diet affected the fatty acid composition in gill and in red blood cells. When transferred to SW, dietary ARA supplementation could promote the bioavailability for eicosanoid synthesis in gill mainly via PLA2g4 activation, and potentially inhibit the stress and inflammatory response caused by different water temperatures through dietary EPA supplementation.

Tailoring freshwater diets towards boosted immunity and pancreas disease infection robustness in Atlantic salmon post smolts.

The aim of the current study was to investigate how freshwater diets impact on immunity in Atlantic salmon smolts in freshwater, during transfer to seawater and in post smolts during the seawater stage with and without pancreas disease (PD) infection. Three specific freshwater diets were prepared: (i) A diet similar in composition to commercial salmon freshwater diets (Standard diet); (ii) A diet composed of vegetable oils (rapeseed, palm and linseed oils) mimicking the fat composition in aquatic insects - the natural diet of wild salmon in freshwater (Fatty acid diet); (iii) A diet enriched with possible immune modulating amino acids including dl-methionine, l-lysine, l-threonine and taurine (Amino acid diet). After seawater transfer, all fish were fed the same commercial diet. Head kidneys were extracted, and their leukocytes isolated from smolts right before transfer to seawater, from post smolts one and six weeks after transfer to seawater, and from post smolts in seawater after 8 weeks of ongoing PD infection. In addition, to provoke bacterial or virus induced inflammation in vitro, the individual leukocyte suspension from all fish were stimulated by lipopolysaccharide (LPS) or polyinosinic acid: polycytidylic acid (PIC). The transfer of smolts from fresh-to seawater changed the transcription of several types of genes. Particularly in isolates from fish fed the Standard or Fatty acid diet in freshwater, overall gene transcription (IL-1ß, CD83, INF-γ, cox2, cd36, MGAT2, catalase) declined. However, the Amino acid diet stimulated the LPS induced gene transcription of IL-1ß, CD83, Cox2, and INF-γ at this stage. In freshwater smolts, PIC stimulated leukocytes showed higher transcription level of Mx and viperin in the Fatty acid and Amino acid diet groups compared to the Standard diet group. In seawater post smolts, Mx and viperin responded similarly to PIC challenge in all diet groups. Furthermore, leukocytes isolated from PD infected fish, continued responding to PIC, regardless of freshwater diet.

Etramp5 as a useful serological marker in children to assess the immediate effects of mass drug campaigns for malaria.

INTRODUCTION: Serological methods provide useful metrics to estimate age-specific period prevalence in settings of low malaria transmission; however, evidence on the use of seropositivity as an endpoint remains scarce in studies to evaluate combinations of malaria control measures, especially in children. This study aims to evaluate the immediate effects of a targeted mass drug administration campaign (tMDA) in Haiti by using serological markers. METHODS: The tMDA was implemented in September-October 2018 using sulfadoxine-pyrimethamine and single low-dose primaquine. A natural quasi-experimental study was designed, using a pretest and posttest in a cohort of 754 randomly selected school children, among which 23% reported having received tMDA. Five antigens were selected as outcomes (MSP1-19, AMA-1, Etramp5 antigen 1, HSP40, and GLURP-R0). Posttest was conducted 2-6 weeks after the intervention. RESULTS: At baseline, there was no statistical difference in seroprevalence between the groups of children that were or were not exposed during the posttest. A lower seroprevalence was observed for markers informative of recent exposure (Etramp5 antigen 1, HSP40, and GLURP-R0). Exposure to tMDA was significantly associated with a 50% reduction in the odds of seropositivity for Etramp5 antigen 1 and a 21% reduction in the odds of seropositivity for MSP119. CONCLUSION: Serological markers can be used to evaluate the effects of interventions against malaria on the risk of infection in settings of low transmission. Antibody responses against Etramp5 antigen 1 in Haitian children were reduced in the 2-6 weeks following a tMDA campaign, confirming its usefulness as a short-term marker in child populations.

Critical evaluation of transcription factor Atf2 as a candidate modulator of alcohol preference in mouse and human populations.

In prior work, congenic strains carrying the DBA/2Igb (D2) region of chromosome 2 (Chr2) for alcohol preference were bred onto a C57BL/6Ibg (B6) background and as predicted were found to reduce voluntary consumption. Subsequently, interval-specific congenic recombinant strains (ISCRS) were generated and also tested. These ISCRS strains reduced the quantitative trait loci (QTL) interval to a comparatively small 3.4 Mb region. Here, we have exploited an integrative approach using both murine and human populations to critically evaluate candidate genes within this region. First, we used bioinformatics tools to search for genes relevant to alcohol preference within the QTL region. Second, we searched for single nucleotide polymorphisms (SNPs) within exons of every gene in this region. Third, we conducted follow-up microarray analyses to identify differentially expressed genes between the B6 and ISCRS strains in mice from each group. Fourth, we analyzed correlations between the expression level of candidate genes and phenotypes of alcohol preference in a large family of BXD recombinant inbred strains derived from B6 and D2. Finally, we evaluated SNP segregation in both BXD mouse strains and in humans who were heavy alcohol drinkers or non-drinkers. Among several potential candidate genes in this region, we identified activating transcription factor 2 (Atf2) as the most plausible gene that would influence alcohol preference. However, the candidacy of Atf2 was only weakly supported when we used a genetic network approach and by focused reanalysis of genome-wide association study data from European-American and African-American populations. Thus, we cannot conclude that Atf2 plays a role in the regulation of the QTL of mouse Chr2.

Dietary beauvericin and enniatin B exposure cause different adverse health effects in farmed Atlantic salmon.

The extensive use of plant ingredients in novel aquafeeds have introduced mycotoxins to the farming of seafood. The emerging enniatin B (ENNB) and beauvericin (BEA) mycotoxins have been found in the novel aquafeeds and farmed fish. Little is known about the potential toxicity of ENNs and BEA in farmed fish and their feed-to-organ transfer. Atlantic salmon (Salmo salar) pre-smolt (75.3 ± 8.10 g) were fed four graded levels of spiked chemical pure ENNB or BEA feeds for three months, in triplicate tanks. Organismal adverse health end-point assessment included intestinal function (protein digestibility), disturbed hematology (red blood cell formation), bone formation (spinal deformity), overall energy use (feed utilization), and lipid oxidative status (vitamin E). Both dietary BEA and ENNB had a low (<∼0.01%) transfer to organs (kidney > liver > brain > muscle), with a higher transfer for ENNB compared to BEA. BEA caused a growth reduction combined with a decreased protein digestion and feed conversion rate- ENNB caused a stunted growth, unrelated to feed utilization capacity. In addition, ENNB caused anemia while BEA gave an oxidative stress response. Lower bench-mark dose regression assessment showed that high background levels of ENNB in commercial salmon feed could pose a risk for animal health, but not in the case of BEA.

The effect of dietary lipid content and stress on egg quality in farmed Atlantic cod Gadus morhua.

The present study assessed differences in fecundity and egg quality from Atlantic cod Gadus morhua fed isoproteic diets containing 13% fat (low fat, LF) or 20% fat (high fat, HF) and either stressed or left unstressed as a control over the spawning season. Each diet was fed to triplicate groups of G. morhua from June 2009, through to first maturation and spawning. In January 2010 sub-groups of G. morhua were moved to land-based spawning tanks where the experimental trial was carried out. At the start of the experiment, G. morhua fed the high-fat diet were significantly larger than G. morhua fed low-fat diet. These differences were maintained through the spawning season, although with a loss of mass in both dietary groups. Relative fecundity through the season was significantly lower in stressed G. morhua fed LF compared to unstressed G. morhua fed the same diet. Stressed G. morhua had a higher variability in weekly amount of eggs spawned, spawning occurred more irregularly, and the spawning period lasted longer than in unstressed G. morhua. Several egg quality variables were also affected: eggs from G. morhua fed LF and exposed to stress had lower fertilization and hatching rates compared to the unstressed G. morhua fed the same diet as well as all G. morhua fed HF. Gadus morhua fed a low-fat diet appeared less tolerant to stress than fish fed a high-fat diet.

Pre-digestion of dietary lipids has only minor effects on absorption, retention and metabolism in larval stages of Atlantic cod (Gadus morhua).

The hypothesis of the present study was that cod larvae have a limitation in lipid digestion, and that absorption of lipids would increase by pre-hydrolysation. The diets used were designed to contain 15% lipid, of which 40% was phosphatidylcholine (PC) and 60 % was TAG. Cod larvae (40d post hatch (dph)) were fed a single meal where either PC or TAG was radioactively labelled, and the labelled PC or TAG was either intact or hydrolysed (pre-digested). The larvae were then incubated individually in chambers with collection of CO2 for 10 h. The following fractions were analysed for radioactivity: the incubation water (evacuated feed); the intestine; the body; the CO2 trap. The larvae ate a 16-29 µg diet, equivalent to 3·4-5·2 % of dry body weight. In the whole population, 0-16% of the lipid was evacuated. The larvae that had eaten less than 1·9-2·7 µg lipid absorbed close to 100% of the lipid, absorption being defined conservatively as the amount contained in the carcass and CO2, excluding the intestinal tissue. In these larvae, approximately 100 % of the absorbed lipid was also catabolised. In the larvae that ingested more than 1·9-2·7 µg lipid, there was a linear reduction in lipid absorption to a minimum of 55% at the highest lipid intakes parallel to an increasing retention of lipids in the carcass. There were only minor differences in digestion, absorption, retention and metabolism of lipids between the larvae fed the different diets, and the larvae tended to retain lipid classes as they were present in the feed. The study shows that 40-dph Atlantic cod larvae have an efficient utilisation of dietary lipids supplied as intact PC and TAG.

Sensitivity and toxic mode of action of dietary organic and inorganic selenium in Atlantic salmon (Salmo salar).

Depending on its chemical form, selenium (Se) is a trace element with a narrow range between requirement and toxicity for most vertebrates. Traditional endpoints of Se toxicity include reduced growth, feed intake, and oxidative stress, while more recent finding describe disturbance in fatty acid synthesis as underlying toxic mechanism. To investigate overall metabolic mode of toxic action, with emphasis on lipid metabolism, a wide scope metabolomics pathway profiling was performed on Atlantic salmon (Salmo salar) (572±7g) that were fed organic and inorganic Se fortified diets. Atlantic salmon were fed a low natural background organic Se diet (0.35mg Se kg-1, wet weight (WW)) fortified with inorganic sodium selenite or organic selenomethionine-yeast (SeMet-yeast) at two levels (∼1-2 or 15mgkg-1, WW), in triplicate for 3 months. Apparent adverse effects were assessed by growth, feed intake, oxidative stress as production of thiobarbituric acid-reactive substances (TBARS) and levels of tocopherols, as well as an overall metabolomic pathway assessment. Fish fed 15mgkg-1 selenite, but not 15mgkg-1 SeMet-yeast, showed reduced feed intake, reduced growth, increased liver TBARS and reduced liver tocopherol. Main metabolic pathways significantly affected by 15mgkg-1 selenite, and to a lesser extent 15mgkg-1 SeMet-yeast, were lipid catabolism, endocannabinoids synthesis, and oxidant/glutathione metabolism. Disturbance in lipid metabolism was reflected by depressed levels of free fatty acids, monoacylglycerols and diacylglycerols as well as endocannabinoids. Specific for selenite was the significant reduction of metabolites in the S-Adenosylmethionine (SAM) pathway, indicating a use of methyl donors that could be allied with excess Se excretion. Dietary Se levels to respectively 1.1 and 2.1mgkg-1 selenite and SeMet-yeast did not affect any of the above mentioned parameters. Apparent toxic mechanisms at higher Se levels (15mgkg-1) included oxidative stress and altered lipid metabolism for both inorganic and organic Se, with higher toxicity for inorganic Se.

The cells and molecules that make a cerebellum.

The molecular underpinnings of cerebellar development are being established through the identification of naturally occurring mutated genes and the knockout of other genes. Sets of genes expressed in the regions of the mes- and metencephalon have been shown to play a crucial role in specifying the cerebellar anlage. Other genes have been shown to be crucial to early granule-cell development, migration of Purkinje and granule cells, and neuron-glia interactions. However, the process of development will ultimately be understood in terms of cellular interactions and the roles that each cell type plays in the assembly of cerebellar structure. One of the most important interactions is between granule and Purkinje cells. This relationship has been shown to be crucial for the control of cell number, migration of neuroblasts and cell differentiation.

Granule cells and cerebellar boundaries: analysis of Unc5h3 mutant chimeras.

Mutations in the Unc5h3 gene, a receptor for the netrin 1 ligand, result in abnormal migrations of both Purkinje and granule cells to regions outside the cerebellum and of granule cells to regions within the cerebellum. Because both Purkinje and granule cells express this molecule, we sought to determine whether one or both of these cell types are the primary target of the mutation. Chimeric mice were made between wild-type ROSA26 transgenic mouse embryos (whose cells express beta-galactosidase) and Unc5h3 mutant embryos. The resulting chimeric brains exhibited a range of phenotypes. Chimeras that had a limited expression of the extracerebellar phenotype (movement of cerebellar cells into the colliculus and midbrain tegmentum) and the intracerebellar phenotype (migration of granule cells into white matter) had a normal-appearing cerebellum, whereas chimeras that had more ectopic cells had attenuated anterior cerebellar lobules. Furthermore, the colonization of colliculus and midbrain tegmentum by cerebellar cells was not equivalent in all chimeras, suggesting different origins for extracerebellar ectopias in these regions. The granule cells of the extracerebellar ectopias were almost entirely derived from Unc5h3/Unc5h3 mutant embryos, whereas the ectopic Purkinje cells were a mixture of both mutant and wild-type cells. Intracerebellar ectopias in the chimera were composed exclusively of mutant granule cells. These findings demonstrate that both inside and outside the cerebellum, the granule cell is the key cell type to demarcate the boundaries of the cerebellum.

Vitamins C and E interact in juvenile Atlantic salmon (Salmo salar, L.).

A two way regression study was performed to investigate the interactions between vitamins C and E, and the influence of dietary vitamin C on the development of vitamin E deficiency in first feeding Atlantic salmon. The fish were fed three levels of all-rac-alpha-tocopheryl acetate (0, 150, and 300 mg/kg), each with six levels of ascorbate monophosphate (0, 7.5, 15, 30, 45, and 60 mg/kg ascorbic acid equivalents). Vitamin C protected the fish against vitamin E deficiency in a dose dependent manner, as seen from the data on growth, mortality, hematology, and lipid oxidation in the liver, indicated by the concentration of malondialdehyde. Vitamin C did not influence the tissue levels of vitamin E, except in vitamin C deficiency, which induced a large drop in liver vitamin E concentration. The liver level of vitamin C increased in response to supplementation of both vitamins. The results indicate two different interaction mechanisms: a synergistic effect of simultaneous protection of the water and lipid phases against oxidation, and regeneration of vitamin E from the vitamin E radical by ascorbic acid.

Analysis of gene action in the meander tail mutant mouse: examination of cerebellar phenotype and mitotic activity of granule cell neuroblasts.

The meander tail (mea) gene results in a stereotypic pattern of cerebellar abnormalities, most notably the virtual depletion of granule cells in the anterior lobe of the cerebellum. The causal basis of this mutation is unknown. In this paper we have taken a three-part approach to the analysis of mea gene action. First, we quantitatively determined the effect of the mea gene on granule cell and Purkinje cell number. We found, in addition to the marked depletion of anterior lobe granule cells ( > 90%), there were also significantly fewer granule cells in the posterior lobe (20-30%) without a concomitant loss of Purkinje cells. Second, we explored the relationship between granule cell depletion caused by the mea gene and by the mitotic poison, 5-fluoro-2'-deoxyuridine (FdU). Prenatal and postnatal ICR mice were treated with FdU to ascertain the regimen that best produces a meander tail-like cerebellar phenotype. The similarity of the effects of the mea gene and injections of FdU at E17 and PO suggests the hypothesis that the mea gene acts to disrupt the cell cycle of cerebellar granule cell precursors. Thus, the third part of this study was to test this hypothesis by using injections of either BrdU (5-bromo-2'-deoxyuridine) or 3H-thymidine into homozygous and heterozygous meander tail littermates at E17 or PO. After processing the tissue for BrdU immunocytochemistry or 3H-thymidine autoradiography, counts were made of the number of labeled and unlabeled external granule layer (EGL) cells to determine the percentage that had incorporated the mitotic label (labeling index). No difference in the labeling index was found between homozygous meander tail mice and normal, heterozygous littermate controls. Therefore, the mitotic activity of the EGL neuroblasts is not disrupted by the mea gene. Furthermore, while a mitotic poison can produce a phenotype similar to the action of the mea gene, mea is phenomenologically different from FdU treatment.

The annexins: specific markers of midline structures and sensory neurons in the developing murine central nervous system.

The annexins are a family of cytoplasmic proteins that have been shown to have numerous actions within a cell. Recent evidence suggests that at least one of these proteins plays a role in the development of the central nervous system (CNS). The present study examines the temporal expression and spatial distribution of annexins I, II, IV, V, and VI during development and at maturity in the murine CNS by immunocytochemical analysis. The results demonstrate that annexins I, II and IV exhibit clear immunolabeling in the murine CNS with distinct patterns of temporal and spatial expression. Annexin IV is the first annexin to be expressed on embryonic day (E) 9.5 while annexin I is the last to be expressed (E11.5). Annexins I, II and IV are found in the floor plate region, but to differing rostrocaudal extents. Annexin I has a very restricted distribution, only present in the midline raphe of the brainstem. Annexin II is present in the spinal cord, brainstem and mesencephalon. Annexin IV has the widest midline distribution, being observed in the floor and roof plates of the developing CNS. Additionally, antibodies against annexin II and IV immunolabel most dorsal root and sensory ganglion cells and their axons. During early postnatal development, immunolabeling with each antibody gradually disappears in many structures, and only first order sensory neurons and their fibers are immunopositive for annexins II and IV at weaning. Three functions of the annexins are suggested by the present findings: (1) to help establish the midline structures of the floor and roof plates, (2) to help direct the decussation of sensory fibers, and (3) to regulate some aspect of sensory neuron processing, such as signal transduction.

Dissociation of spatial navigation and visual guidance performance in Purkinje cell degeneration (pcd) mutant mice.

Spatial learning in rodents requires normal functioning of hippocampal and cortical structures. Recent data suggest that the cerebellum may also be essential. Neurological mutant mice with dysgenesis of the cerebellum provide useful models to examine the effects of abnormal cerebellar function. Mice with one such mutation, Purkinje cell degeneration (pcd), in which Purkinje cells degenerate between the third and fourth postnatal weeks, were evaluated for performance of spatial navigation learning and visual guidance learning in the Morris maze swim-escape task. Unaffected littermates and C57BL/6J mice served as controls. Separate groups of pcd and control mice were tested at 30, 50 and 110 days of age. At all ages, pcd mice had severe deficits in distal-cue (spatial) navigation, failing to decrease path lengths over training and failing to express appropriate spatial biases on probe trials. On the proximal-cue (visual guidance) task, whenever performance differences between groups did occur, they were limited to the initial trials. The ability of the pcd mice to perform the proximal-cue but not the distal-cue task indicates that the massive spatial navigation deficit was not due simply to motor dysfunction. Histological evaluations confirmed that the pcd mutation resulted in Purkinje cell loss without significant depletion of cells in the hippocampal formation. These data provide further evidence that the cerebellum is vital for the expression of behavior directed by spatial cognitive processes.

Effects of alcohol exposure during different periods of development: changes in hippocampal mossy fibers.

Exposure to 10-12 g/kg/day of alcohol either during days 1-10 or 11-21 of gestation had no detectable effect on hippocampal mossy fiber development. Exposing artificially reared rat pups to 7.0-7.5 g/kg/day of alcohol during days 1-10 postpartum dramatically altered the organization of the Timm-stained mossy fiber terminal field when the animals were examined as adults, suggesting that alcohol exposure during a period equivalent to the human third trimester is more deleterious to brain development than exposure during periods equivalent to either the first or second trimesters.

Differential strain susceptibility following 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) administration acts in an autosomal dominant fashion: quantitative analysis in seven strains of Mus musculus.

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) has been used as a potent neurotoxin to approximate, in animals, the pathology that is observed in human Parkinson's disease. In this study, we examine the toxicity of MPTP in seven strains of mice, spanning a genetic continuum of Mus musculus as a prelude to uncovering complex traits associated with MPTP toxicity. Seven days following injection of 80 mg/kg MPTP (4x20 mg/kg every 2 h), we find that the individual mouse strains exhibit dramatic differences in SNpc neuron survival, ranging from 63% cell loss in C57BL/6J mice to 14% cell loss in Swiss-Webster (SW) mice. In order to determine if the susceptibility trait was dominant, additive or recessive, we crossed C57Bl/6J mice with either SWR/J or AKR/J mice and examined the effect of MPTP on F1 C57BL/6JxSWR/J or F1 C57BL/6JxAKR/J animals. We find that all of the F1 animals were phenotypically identical to the C57BL/6J animals. In addition, no gender differences were noted in any of the MPTP-treated inbred mice or in the F1 animals. These results suggest that susceptibility to cell loss following MPTP is autosomal dominant and this polymorphism is carried on the C57BL/6J allele.

Regional differences in the timing of dendritic outgrowth of Purkinje cells in the vermal cerebellum demonstrated by MAP2 immunocytochemistry.

Detailed, within-subjects Golgi analyses of regional differences in cerebellar Purkinje cell dendritic development are impractical due to the capriciousness of that technique. Immunocytochemical labeling of microtubule-associated protein 2 (MAP2) was used to reveal the dendritic development of Purkinje cells, and indicated marked differences in the timing of initial outgrowth of Purkinje cell dendrites for different lobules in the developing rat cerebellar vermis. In particular, an early maturing region of Purkinje cell dendritic outgrowth (lobules I, II, IX and X and along the primary fissure), and a late maturing region (distal lobule VI, lobule VII and dorsal lobule VIII) were documented.

The development of laminar staining for neuron-specific enolase in the rat somatosensory cortex.

The expression of the enzyme neuron-specific enolase (NSE) in the central nervous system (CNS) has been used as a developmental marker based on observations that it is expressed shortly after the arrival of afferent inputs. The immunostaining pattern of NSE was examined in the laminae of the somatosensory cortex of the rat and the relationship of this staining pattern with previous data on the timing of afferent and efferent arrival was determined. Male rat pups were sacrificed on postnatal days 1 (24 h after birth), 3, 5, 8, 10, 12, 15 and 20, and as an adult (over 90 days of age). Sections were stained with an anti-NSE antibody using the avidin-biotin immunocytochemical method. Sections from day 1 animals revealed stained cells in the subplate layer and cortical plate, presumably in cells destined to form layers VI and V. By day 8 there was staining in layers II, III, V and VI, the same layers that exhibited staining in the adult rat. This appears consistent with the arrival of afferents and efferents which is completed by approximately postnatal day 7. On day 10, there was a change in the staining pattern: cell staining in layer VI was decreased and then increased gradually up to adult levels by day 20. A stable pattern of NSE staining was not observed previous to day 20. These results suggest that changes in NSE expression following the initial arrival of afferents may relate to maturation of the neurons.