5th Postnatal Day Research Articles

Changes in Growth and Pancreatic mRNA Concentrations During Postnatal Development of Rat Pancreas

Changes in pancreatic growth and in mRNA concentrations in rat pancreas were monitored by dot-blot hybridization with cloned cDNAs of rat amylase, chymotrypsinogen B, proinsulin I, and actin during the pre- and postnatal period in the rat. Wistar rats were killed at the 18th day of gestation and at the 1st, 10th, 20th, 35th, and 87th day of postnatal life. It was concluded from the ratio of pancreatic weight/body weight that pancreatic growth preceded body growth. Pancreatic protein and total RNA concentration increased 2.9 times during the period studies. All studied mRNAs increased in concentration during the postnatal development period. Messenger RNA for chymotrypsinogen B and proinsulin I exhibited a significant increase after birth, decreased by the 10th day of life, and increased thereafter. For amylase mRNA, no significant changes were observed around birth, a progressive increase occurring thereafter up to the 87th day of life. The mRNA for actin showed a progressive increase between the 18th day of gestation and the 20th postnatal day, after which it remained stable. We concluded that each mRNA showed a singular profile of increase during postnatal development.

Relation between axons and oligodendroglial cells during initial myelination I. The glial unit

The morphology of oligodendroglial-axon units was examined by electron microscopy during ensheathment and initial myelination in developing feline spinal cord and corpus callosum white matter. In addition to a qualitative examination of single sections from many stages of development, a morphological analysis of spinal cord and corpus callosum units was made on the basis of serial sections from a few stages. The results show that myelination commences around embryonic/fetal day 40 and the 20th postnatal day in the spinal cord and corpus callosum areas, respectively. In both areas immature glial cells, lacking the cytological features of typical oligodendrocytes, initially associate with several axons and provide them with cytoplasmic sheaths. Serial section analysis of units, which have begun formation of compact myelin, indicates that individual cells are associated with single myelin sheaths in the spinal cord area, in a way principally similar to the Schwann cell-myelin units in developing peripheral nerves. This suggests the possibility that early spinal cord oligodendrocytes might shift from a polyaxonal to a monoaxonal association after initial ensheathment and before formation of compact myelin. In the corpus callosum area the examined serially-sectioned cells were found to be connected to several myelin sheaths through long thin processes. The myelin sheaths related to one cell are relatively uniform in terms of number of myelin lamellae and axon diameter, but the clockwise/counter-clockwise course of the myelin spiral varies randomly. Units containing both homogeneously uncompacted (cytoplasmic) and fully compacted (myelin) sheaths have not been found. In both areas the ensheathing cells achieve an oligodendrocyte-like cytology during formation of the first layers of compact myelin. These observations support the view that oligodendrocytes are structurally heterogeneous: those myelinating prospective large axons seems to differ from those myelinating axons destined to remain small. The possible functional and pathophysiological implications of this heterogeneity remain to be elucidated.

Lectin staining of saccharides in the normal and hypothyroid developing organ of Corti

Lectin staining has been used to detect mono- and oligosaccharides in normal and hypothyroid developing organs of Corti in the rat. Eight developmental stages were studied (1, 5, 8, 10, 15, 20, 50 and 60 days after birth). Congenital hypothyroidism was induced by oral administration of propylthyouracil to pregnant rats. Labelling of the tectorial membrane with 3 lectins, Ulex europaeus agglutinin-I (UEA-I), Lens culinaris agglutinin (LCA) and Ricinus communis agglutinin-I (RCA-I) showed no significant differences between normal and hypothyroid animals. Staining with peanut agglutinin (PNA) showed that the hypothyroid adult tectorial membrane (but not the normal one) possesses the disaccharide galactose+ N-acetyl galactosamine. Phaseolus vulgaris agglutinin-L (PHA-L) labels the whole tectorial membrane in both groups of animals, but the staining is more intense in the hypothyroid one for a narrow band of oligosaccharide located just between the tectorial membrane and the underlying organ of Kölliker. Both soybean agglutinin (SBA) and succinylated wheat germ agglutinin (WGA) stain the tectorial membrane as well as the cytoplasm of the cells constituting the inner portion of the organ of Kölliker; this latter feature disappears in the normal animals about the 8th postnatal day, but it is abnormally preserved until the 60th postnatal day in the hypothyroid ones. In the adult hypothyroid animals, 3 of the lectins (LCA, PHA-L and WGA) stain extracellular conglomerates located under the synaptic pole of the outer hair cells.

Application of analytical color fluorescence electron microscopy to biomedical field: I. Vitamin A ester in rat retina.

An analytical color fluorescence electron microscope (ACFEM) based on a high-resolution scanning electron microscope has been developed. The ACFEM enables us not only to detect cathodoluminescence (CL), which is a weak luminescence signal under electron beam bombardments, as color images, but also to analyze CL spectra. A cryo-SEM method was introduced to prevent beam effect on biological specimens. In experiments 1 and 2, we observed adult rat retinas under different conditions: hyper- and hypovitaminosis A and light and dark adaptation, which revealed that the distribution of vitamin A ester and its change under these conditions could be detected in situ by the ACFEM. In experiment 3, postnatal development of rat retina was observed under the ACFEM up to 3 weeks after birth. The retinal pigment epithelial cells of new born rats were already functioning as vitamin A storing cells. On the other hand, vitamin A ester in the developing outer segment first appeared on the 13th postnatal day, which suggests a correlation to the development of visual function. These results show that CL analysis by the ACFEM is a simple and effective new method in the field of histo- and cytochemistry.

Frequency selectivity in the auditory periphery: Similarities between damaged and developing ears

Single fiber tuning curves (stimulus frequency versus neural threshold curves) were obtained from 198 auditory nerve fibers in 24 kittens between birth and the 16th postnatal day and from 74 auditory nerve fibers in adult cats. Three developmental stages during which adult-like frequency-resolving capacity was acquired were identified. During the early stage of postnatal development, all auditory nerve fibers were essentially untuned and responded to a narrow range of low to middle frequency tone bursts presented at intensities exceeding 110 dB sound pressure level (SPL) re 20 μPa. In the intermediate stage, which occurred during the second postnatal week, auditory nerve fibers tuned to low- and mid-range frequencies acquired adult-like frequency-resolving capacity. Fibers tuned to high frequencies, which were recorded later in development than those tuned to lower frequencies, were as sharply tuned as their adult counterparts, but exhibited a low contrast between thresholds at characteristic frequency (tip) and lower (tail) frequencies (ie, low tip-to-tail ratios). Adult-like tuning curves were observed during the third stage, primarily as a consequence of the acquisition of adult-like tip-to-tail ratios. Our understanding of the cochlear mechanism(s) by which frequency selectivity is produced in adult animals has recently been enhanced by a combined anatomy and physiology investigation conducted by Liberman and Dodds, 1 in which clear anatomic foci of cochlear damage were identified in cats with functionally characterized hearing loss. Similarly, descriptions of anatomic differentiation in the feline auditory end-organ 2 correlate with functional measures of peripheral auditory system development. In this report, anatomic and physiologic similarities between developing and damaged ears are considered in an attempt to better characterize the process whereby normal frequency selectivities and thresholds are developed. Our findings support the notion that anatomic changes in the cochlea during development, primarily the development of adult-like anatomic relations between the tectorial membrane and sensory cells, underlie the acquisition of adult-like auditory nerve fiber tuning.

The effect of hypothyroidism on the development of the glycogen content of organ of Corti's hair cells

The density of glycogen particles in organ of Corti's sensory cells was measured to determine the effect of congenital hypothyroidism upon the normal development of this energy source. This density in both normal and hypothyroid inner hair cells remains in low values from birth to adulthood. On the other hand, that of normal outer hair cells undergoes a great increase between the 10th and the 20th postnatal days, coinciding with the maturation of both the efferent innervation of these cells and the tuning properties of the auditory receptor. The glycogen stores of the hypothyroid outer hair cells do not show any significant increase from birth to adulthood. This latter fact suggests that the congenital hypothyroidism restrains the development of an important energy source of outer hair cells, most surely disturbing the physiological processes relying on glycogen metabolism.

Quantitative electron microscopic analysis of postnatal development of zinc-positive nerve endings in the rat amygdala using Timm's sulphide silver technique

The distribution and quantitation of zinc during postnatal development of the rat amygdala were investigated by light and electron microscopy with Timm's sulphide silver method. The adult rat amygdala could be divided into basolateral, basomedial, central, cortical, intercalated, lateral and medial subnuclei on the basis of Timm's plus Toluidine blue staining. Only a very weakly positive Timm's reaction could be observed in newborns, and a positive reaction was seen on the 5th postnatal day. The reaction became stronger with development, and reached adult levels by the 30th postnatal day. Electron microscopically, we investigated the basolateral subnucleus of the amygdala. The electron-dense deposits of silver grains from the Timm's reaction were only seen in nerve fibers endings containing many small clear vesicles. The results are similar to those described for mossy fiber endings in the rat hippocampus and in boutons of other telencephalic structures. The number of deposits of silver grains increased with age, and reached that of the adult between the 20th and 30th postnatal days. A significant increase in the number of silver grains per μ 2 area of the Timm's-positive nerve terminals occurred between the 10th and 20th postnatal days. Based on the available literature, these findings suggest that zinc exists in nerve fiber endings in the rat amygdala and that the amount increases with postnatal age. The findings are consistent with an important role for zinc in synaptic transmission.

Evolution of Some Biochemical Characteristics of the Intestinal Mucosa During the First Postnatal Weeks in C57 Mice. Effects of Thyroxine and Putrescine

In C57 mice, during the third week after birth, there is a rapid conversion of the intestinal epithelium from fetal to mature adult status. Some characteristics (lactase, maltase, diamine oxidase, and sucrase activity; putrescine, spermine and spermidine concentrations; mitotic index) have been analyzed for the proximal, middle, and distal parts of the intestine in mice of different ages: 8, 15, 19, and 60 days. The most important observations recorded were as follows: (a) sucrase- and maltase-specific activities as well as spermine and spermidine contents increased abruptly on the 19th postnatal day and then decreased; (b) decrease of lactase and diamine oxidase-specific activity was recorded between the 15th and 19th postnatal days, (c) later, diamine oxidase-specific activity increased, whereas putrescine content decreased in the proximal part of the intestine; and (d) mitotic index was not significantly different when estimated for the crypts on days 11 and 19. No similar variations of these biochemical parameters were observed over a period of 3 days when mice were injected with thyroxine on the 8th day or when mice received putrescine per os on the 9th day, as explained in the text. Only a slight but significant variation in sucrase- or maltase-specific activity with thyroxine and a variation of the lactase or DAO-specific activity of the distal part of the intestine with putrescine were recorded.

Evolution of Some Biochemical Characteristics of the Intestinal Mucosa During the First Postnatal Weeks in C57 Mice. Effects of Thyroxine and Putrescine

In C57 mice, during the third week after birth, there is a rapid conversion of the intestinal epithelium from fetal to mature adult status. Some characteristics (lactase, maltase, diamine oxidase, and sucrase activity; putrescine, spermine and spermidine concentrations; mitotic index) have been analyzed for the proximal, middle, and distal parts of the intestine in mice of different ages: 8, 15, 19, and 60 days. The most important observations recorded were as follows: (a) sucrase‐ and maltase‐specific activities as well as spermine and spermidine contents increased abruptly on the 19th postnatal day and then decreased; (b) decrease of lactase and diamine oxidase‐specific activity was recorded between the 15th and 19th postnatal days, (c) later, diamine oxidase‐specific activity increased, whereas putrescine content decreased in the proximal part of the intestine; and (d) mitotic index was not significantly different when estimated for the crypts on days 11 and 19. No similar variations of these biochemical parameters were observed over a period of 3 days when mice were injected with thyroxine on the 8th day or when mice received putrescine per os on the 9th day, as explained in the text. Only a slight but significant variation in sucrase‐ or maltase‐specific activity with thyroxine and a variation of the lactase or DAO‐specific activity of the distal part of the intestine with putrescine were recorded.

Postnatal ontogeny of POMC gene expression in the rat pituitary: an analysis by in situ hybridization histochemistry

Postnatal development of proopiomelanocortin gene expression in the rat pituitary was examined using in situ hybridization histochemistry. In adult rats, a very high density of hybridization signals was seen in the intermediate lobe of the pituitary, while only a moderate density occurred in the anterior lobe. No evidence of hybridization was detected in the posterior lobe. At birth, both the intermediate and anterior lobes had low to moderate frequencies of hybridization signals but a rapid rise to moderate density was noted by the 8th postnatal day. Radioactive labelling in the intermediate lobe continued to increase sharply with age to reach a plateau at postnatal day 28, while hybridization signals in the anterior lobe levelled off at postnatal day 8 with no subsequent rise in density.

The fatty liver dystrophy (fld) mutation: A new mutant mouse with a developmental abnormality in triglyceride metabolism and associated tissue-specific defects in lipoprotein lipase and hepatic lipase activities

An autosomal recessive mutation, termed fatty liver dystrophy (fld), can be identified in neonatal mice by their enlarged and fatty liver (Sweet, H. O., Birkenmeier, E. H., and Davisson, M. T. (1988) Mouse News Letter 81, 69). We have examined the underlying metabolic abnormalities in fld/fld mice from postnatal days 3-40. Serum and hepatic triglyceride levels were elevated 5-fold in suckling fld/fld mice compared to their +/? littermates but abruptly resolved at the suckling/weaning transition. Blot hybridization analysis of liver and intestinal RNAs revealed a liver-specific increase in apolipoprotein (apo) A-IV and C-II mRNA concentrations (100- and 6-fold, respectively) that was limited to the suckling and early weaning stages in fld/fld mice. Resolution of these differences during the weaning period could not be delayed by prolonging suckling to the 20th postnatal day nor could the mutant phenotype be elicited in young adult animals with a high fat diet. Lipoprotein lipase (LPL) activity was reduced 16-fold in the white adipose tissue of fld/fld mice until the onset of weaning. Heart activity was decreased less than 2-fold, but there were no deficits in brown adipose tissue or liver. Hepatic lipase (HL) mRNA levels and activity were significantly reduced in fld/fld livers and sera, respectively, during the suckling period. Mapping studies show the fld locus to be distinct from loci encoding LPL, HL, and apoA-IV, and those responsible for the combined lipase deficiencies in cld/cld and W/Wv mice. These data suggest that the fld mutation is associated with developmentally programmed tissue-specific defects in the neonatal expression of LPL and HL activities and provide evidence for a new regulatory locus which affects these lipase activities. This mutation could serve as a useful model for (i) analyzing the homeostatic mechanisms controlling lipid metabolism in newborn mice and (ii) understanding and treating certain inborn errors in human triglyceride metabolism.

Postnatal development of the Sertoli cell barrier, tubular lumen, and cytoskeleton of Sertoli and myoid cells in the rat, and their relationship to tubular fluid secretion and flow.

The postnatal development of the Sertoli cell barrier, tubular lumen, fluid flow, and cytoskeletal elements in Sertoli and myoid cells was investigated in the Sprague-Dawley rat. With the aid of hypertonic fixatives, a barrier to the rapid entry of fluid was noted in the majority of tubules on the 15th and 16th postnatal (p.n.) days and was completely formed in all tubules prior to p.n. day 18. The actin forming the ectoplasmic specialization (ES), a cytoskeletal complex related to the occluding junctions composing the barrier, began its development during the period of initial barrier formation (16 p.n. day) and progressively attained its adult prominence. The ES developed its characteristic adult pattern and adult fluorescent intensity at about p.n. day 22. Some seminiferous tubules showed very small lumina as early as p.n. day 10. All tubules were not open until p.n. day 30. The size (diameter) of the lumen increased slowly from p.n. day 10 until p.n. day 30 when it started to increase rapidly until about p.n. day 50. Fluid flow in seminiferous tubules was detected as early as p.n. day 20 and increased in amount thereafter. Myoid cell actin filament bundles, running in parallel, were present at p.n. day 10. Actin formed a meshwork pattern characteristic of the adult on, or slightly prior to, p.n. day 22. These data indicate that there is a temporal relationship between the development of the actin cytoskeleton within the Sertoli cell and initial formation of the Sertoli cell barrier. Similarly, there is a temporal relationship between the development of the actin cytoskeleton of myoid cells and tubular fluid flow. The rapid increase in tubular lumen diameter, however, does not correlate with the initial development of Sertoli and myoid cytoskeletal elements.

Effect of maternal exposure to smoke on gas diffusion capacity in neonatal rat

Offspring of control and experimental (chronically exposed to whole cigarette smoke) rats were sacrificed on 15th postnatal day. Pulmonary tissues were processed for quantitative electron microscopic analyses. Arithmetic and harmonic mean thicknesses were directly measured and diffusion capacity was derived for separate layers (tissue, plasma, erythrocyte) and the entire lung. Experimental offspring showed a greater volume density of interstitium in parenchyma ( P < 0.04), arithmetic ( P < 0.05) and harmonic ( P < 0.009) mean thicknesses. Diffusion capacities for lungs of both animal groups, however, were similar (0.00543 and 0.0054 cm 3 O 2 · min −1 · mm Hg −1 · g body weight −1 for control and experimental lungs respectively). Experimental tissue appeared better adapted for optimal exchange of gases ( P < 0.05), as indicated by corrugation index (ratio between arithmetic and harmonic mean thicknesses) than for the control counterpart. In spite of induced alterations in several developmental processes by a presently used experimental procedure, tissue capacity for transport of gases was not significantly changed in either the alveolo-capillary membrane or the entire lung.

Ontogeny of tyrosine hydroxylase-positive but dopamine β-hydroxylase-negative neuron-like cells in the pineal gland of golden hamsters

The ontogeny of tyrosine hydroxylase (TH)-positive but dopamine β-hydroxylase (DBH)-negative neuron-like cells in the pineal gland of golden hamsters was investigated by double immunostaining. These cells first appeared on the 6th postnatal day and thereafter increased in number. On the other hand, TH- and DBH-positive nerve fibers, probably originating from the superior cervical ganglion, were already present in the pineal gland at birth.

Location of a transiently expressed glycoprotein in developing cerebellum delineating its possible ontogenetic roles

The development pattern of a 31,000 mol. wt phosphatidyl inositol-anchored membrane glycoprotein was followed during development in mouse and rat cerebellum using monoclonal antibody 194–653. The epitope was developmentally regulated and particularly abundant in post mitotic precursors of granule cells, newly formed parallel fibres and unmyelinated axons of the white matter between the 5th and the 15th postnatal days. It decreased considerably thereafter. In the adult, a significant although relatively low staining was observed only in white matter. Observation at the ultrastructural level showed that most of the 31,000 mol. wt glycoprotein was very concentrated on neuronal plasma membranes. A little immunoreactivity was also found intracellularly at the perinuclear membrane of neuroblasts of the external germinal layer. The antigen was present in the coated pits and intracellularly in coated vesicles. Immunochemical studies indicated that 31,000 mol. wt antigen was very likely to be a previously identified transient concanavalin A-binding glycoprotein insoluble in neutral detergents ( Reeber et al., 1981 ; Brain Res. 229, 53–65). It appeared to be one of the glycoprotein ligands for two endogenous mannosyl-lectins isolated from rat cerebellum ( Zanetta et al., 1985 , Devl. Brain Res. 17, 233–243, Zanetta et al., 1987 , J. Neurochem. 49, 1250–1257). The affinity of the 31,000 mol. wt glycoprotein for the two endogenous lectins, together with its developmental pattern and localization indicate that it could be an important molecule for contact guidance during migration of neurons and for myelination and could take part in other ontogenetic steps.

Early postnatal treatment with propranolol affects development of brain amines and behavior.

The present study examined the effects of early postnatal treatment with a beta-adrenoceptor antagonist propranolol (5 mg/kg IP daily) on concomitant and subsequent behavior and central aminergic transmission in rats. During propranolol exposure from the 7th to the 20th postnatal days sleep-wake recordings, carried out with the static charge sensitive bed (SCSB) method, showed a decrease in the percentage of active sleep and an increase in waking. When the animals were 1-3 months of age, the open field behavior was changed, immobility time in the Porsolt's swim test was lengthened, and voluntary alcohol consumption was increased in the propranolol-treated rats. Neither motor reactivity to auditory stimuli nor spontaneous alternation behavior was affected. At the age of 4 months concentrations of brain amines and their metabolites were measured from several brain regions. In the propranolol-treated rats the noradrenaline levels were increased in the limbic forebrain and cerebellum. The results suggest that in rats the exposure to propranolol during the rapid growth period of cerebral catecholamine systems, and the concomitant alterations in sleep are related to later changes in behavior and to increased noradrenaline content in the limbic forebrain and cerebellum.

Spermine and spermidine induce intestinal maturation in the rat

In the present study, we aimed to induce precocious intestinal maturation in neonatal rats by the oral administration of polyamines. Groups of 5 rats received either saline, spermidine (10 μmol daily), or spermine (6 μmol daily) orally on the 12th, 13th, and 14th postnatal days. The rats were killed on the 15th postnatal day. After the small bowel was removed, a 1-cm distal ileal segment was removed for histologic examination and the remaining small bowel tissue was homogenized for further biochemical analysis. Polyamine administration was shown to induce structural and biochemical mucosal changes characteristic of postnatal maturation. Lactase, sucrase, and maltase specific activities (micromoles of substrate hydrolyzed per minute per gram of protein) were 80 ± 10, 10 ± 3, and 116 ± 19 for the saline-treated rats; 51 ± 7, 34 ± 2, and 315 ± 37 for the spermidine-treated rats; and 25 ± 2, 46 ± 5, and 419 ± 63 for the spermine-treated rats, respectively. Similar results were obtained with rats, first treated with spermine (6 μmol) on the 7th postnatal day, receiving spermine (6 μmol) daily as described above and killed on the 10th postnatal day. Dose-response experiments performed as reported above in rats whose treatment began on the 12th postnatal day showed that the maturational effects of orally administered spermine are dose-dependent.

The twitcher mouse: accumulation of galactosylsphingosine and pathology of the sciatic nerve

Morphological and biochemical changes were investigated in the early developmental stages of sciatic nerve of the twitcher mouse, a murine model of human globoid cell leukodystrophy. The concentration of galattosylsphingosine (psychosine) and the chromological changes of the twitcher mouse peripheral nerve pathology correlated well. Galactosylsphingosine had already accumulated at birth and dramatically increased with age. Characteristic inclusions were observed in Schwann cells and macrophages of the twitcher mouse on the 5th postnatal day. Endoneurial edema developed after 10 postnatal days and the hypomyelination was pronounced at 15–20 postnatal days. These findings suggest that galactosylsphingosine is cytotoxic for myelin-forming cells and is closely related to pathogenetic events in the twitcher mouse.

Ontogenesis of peptidylglycyl α-amidation activity in the mouse hypothalamus in vivo and in serum-free medium cultures. Relation with thyroliberin (TRH) accumulation and release in vitro

The influence of peptidylglycine α-amidating monooxygenase (PAM) and of its co-factor, ascorbate, were studied in relation to thyroliberin (TRH) activity during mouse hypothalamus development. In vivo, PAM activity developed slowly at fetal stages, and exhibited a sharp rise around the 5th–8th postnatal day, the adult level being reached around day 15. The same developmental pattern was observed when studied in serum-free cultures initiated from fetal mouse hypothalamus. Using this in vitro model, we investigated the effects of ascorbate, a necessary co-factor of PAM, on TRH. Upon ascorbate supplementation of the culture medium, the TRH accumulation normally occurring in our cultures was further enhanced. The half maximum effect was attained with 20 μM, and the amplitude of the response to ascorbate was maximum around 9–13 days in vitro. Moreover, ascorbate increased to an even larger extent the amounts of TRH released upon chemical depolarization. These results are consistent with a direct role of ascorbate on PAM activity, but other more general effects on the maturation of the neuronal response to physiological stimuli cannot be excluded.

An increase in striatal Met-enkephalin-like immunoreactivity in neonatally dopamine-depleted rats

A pronounced decrease in dopamine content in both the striatum and the mesolimbic area was observed up to 120 days of age after treatment with 6-hydroxydopamine (100 μg, intracisternally) following desipramine pretreatment (20 mg/kg, i.p.) at the 5th postnatal day. In the 6-hydroxydopamine-treated rats, Met-enkephalin-like immunoreactivity in the striatum was found to have increased at 35 and 70 days old, although it remained unchanged at 25 days old. The neonatal administration of 6-hydroxydopamine, however, did not affect the peptide content in the mesolimbic area at any age. These results indicate that the neonatal dopamine reduction changes the Met-enkephalinergic neuronal activity in the striatum at the adult age.