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D A Carter

Publications and source records attributed to D A Carter.

At least 19 recordsLinked to original sources
PubMed1992

The identification of a cis-acting element involved in cyclic 3',5'-adenosine monophosphate regulation of bovine vasopressin gene expression.

Cyclic adenosine 3',5'-monophosphate (cAMP) has been implicated as an intracellular messenger mediating osmotic regulation of expression of the gene encoding the neuropeptide vasopressin (VP) in the hypothalamus. We have used a heterologous transient transfection system to demonstrate that cAMP regulates the bovine VP gene promoter following transfection into CV1 cells. Mutational analysis identified a bovine VP cAMP-responsive element (BVP-CRE) 120-112 base-pairs upstream of the start of transcription. DNase I footprint analysis using nuclear protein extract from CV1 cells showed protection at the site of the BVP-CRE. Protection of the BVP-CRE was also observed using purified AP1 protein, while there was a weak interaction with the BVP-CRE using purified rat CREB protein. Nuclear proteins purified from the rat supraoptic nucleus bind to the BVP-CRE. As transgenic mouse studies have shown that the bovine VP gene is subject to appropriate physiological regulation in the mouse hypothalamus (Ang, H. L., Funkhouser, J., Carter, D. A., Ho, M. Y., and Murphy, D. (1991) Soc. Neurosci. Abstr. 513, 12), these data indicate a role for the BVP-CRE element in mediating VP gene expression in vivo. These data demonstrate that cAMP regulates bovine VP gene expression in vitro via a cis-acting element within the VP promoter, and this activation may be mediated by members of the AP1/ATF/CREB family of transcription factors.

Animals↗
PubMed1992

3' Alu PCR: a simple and rapid method to isolate human polymorphic markers.

Microsatellites, such as (TG)n found at random throughout the genome, or as 3' extensions of Alu sequences are being increasingly used as genetic markers because of their pluriallelic character. The search for polymorphic microsatellites is time consuming, however, as it is necessary to sequence clones containing the microsatellites sequences in order to design specific PCR primers before testing for polymorphism, which does not always occur. We propose here a new approach to generate polymorphic markers, based on the amplification of microsatellites at the 3' end of Alu sequences, without the need for cloning or sequencing steps.

Base Sequence↗
PubMed1992

Decrease in hypothalamic vasopressin mRNA poly(A) tail length following physiological stimulation.

1. The vasopressin mRNA in the adult male rat hypothalamus is modulated in two distinct ways by a dehydration stimulus. In addition to the well-established increase in transcript abundance, it has recently been demonstrated that the vasopressin mRNA poly(A) tail increases in length. 2. We have studied the ontogeny of poly(A) tail length modulation in neonates in response to milk deprivation and found that poly(A) tail length changes are age dependent. In neonates older than 12 days of age, the vasopressin mRNA poly(A) tail length increased with milk deprivation and this effect became more marked in older animals. However, in rats 5 to 9 days old, milk deprivation resulted in a detectable though not significant decrease in vasopressin mRNA poly(A) tail length. 3. As milk deprivation is a combination of dehydration and starvation, we investigated the effect of the latter stimulus in more mature animals. We found that starvation modifies the length of the vasopressin mRNA poly(A) tail in a manner opposite that due to dehydration. 4. Our data indicate a novel mode of regulation of the vasopressin mRNA, namely, poly(A) tail shortening. This system provides a model for future studies concerning the adaptive role of poly(A) tail length modulation in response to physiological stimuli.

Adaptation, Physiological↗
PubMed1992

Nuclear mechanisms mediate rhythmic changes in vasopressin mRNA expression in the rat suprachiasmatic nucleus.

Vasopressin (VP) gene expression in the rat suprachiasmatic nucleus (SCN) is subject to a cyclical mode of regulation which is indicative of a close association with the circadian clock intrinsic to this area of the hypothalamus. Previous studies show that both the amount and size (due to differential polyadenylation) of VP mRNA are reduced during the dark phase of the daily cycle. We have now identified the cellular site wherein these changes are mediated. By transcriptional run-on analysis of nuclei isolated at different time points from the SCN we have shown that an attenuation of transcriptional activity can account for the dark-phase reduction in VP mRNA levels; by comparison with other genes expressed in this tissue, a significant, VP gene-specific reduction was observed which resulted in dark-phase transcriptional activity at 30% of light-phase activity (P less than 0.005). A similar diurnal variation was not found in the supraoptic nucleus. In addition, by Northern analysis of sub-cellular RNA pools, we have demonstrated that the smaller, dark-phase-specific VP RNA species is located, in abundance, within the nuclear fraction. These results provide clear evidence that the cyclical changes in SCN VP mRNA expression are primarily regulated within the nucleus, indicating that any potential regulation in the cytoplasm is of secondary importance. Further analysis of the molecular components which mediate the cyclical changes in transcriptional activity of the VP gene may identify fundamental aspects of neuronal timing mechanisms.

Animals↗
PubMed1992

Neurotransmitter-stimulated immediate-early gene responses are organized through differential post-synaptic receptor mechanisms.

The products of the cellular immediate-early genes (IEGs) are thought to act as messengers in the coupling of trans-synaptic stimuli with altered neuronal gene expression. However, the manner in which neurotransmission specifies particular responses through the IEGs is undefined. In this report, mRNA and transcription analysis of a precisely-timed, physiological IEG response illustrates how an IEG signal may be organized through differential neurotransmitter receptor activation. The nocturnal pattern of IEG expression in the rat pineal gland has been shown to be differentially regulated through post-synaptic adrenergic receptors. Induction of the c-fos gene is primarily mediated through alpha 1-receptors, whereas the coordinately regulated jun-B gene exhibits dual regulation through alpha 1- and beta-receptors. A simultaneous repression of c-jun expression is partly mediated through a beta-receptor mechanism. In vitro analysis of IEGs in cultured pineal glands has confirmed the receptor-specific link between adrenergic neurotransmission and IEG induction. The pineal is a unique neuroendocrine model in which the characteristics and function of the IEG third messenger system may be defined.

Animals↗
PubMed1991

Retinal ganglion cell terminals in the hamster superior colliculus: an ultrastructural study.

The distribution, cross-sectional area, and presynaptic and postsynaptic characteristics of retinal ganglion cell axon terminals in the superior colliculus of normal adult female Syrian hamsters were investigated by quantitative ultrastructural techniques. After an intravitreal injection of horseradish peroxidase, most labelled axon terminals were found in the stratum griseum superficiale and stratum opticum of the contralateral superior colliculus. However, a small proportion (approximately 2%) of retinal ganglion cell axon terminals were located in deeper layers of the superior colliculus between the stratum opticum and the periaqueductal grey matter. Terminals were smaller in the upper two-thirds of the stratum griseum superficiale than in the lower one-third of this layer, the stratum opticum, and the stratum griseum intermedium. Presynaptic characteristics such as the length and number of contacts and the postsynaptic neuronal domains (somata, dendritic spines, or shafts) contacted by retinal ganglion cell axons in the superior colliculus were similar in all layers.

Animals↗
PubMed1991

Degenerative and regenerative responses of injured neurons in the central nervous system of adult mammals.

In adult mammals, the severing of the optic nerve near the eye is followed by a loss of retinal ganglion cells (RGCs) and a failure of axons to regrow into the brain. Experimental manipulations of the non-neuronal environment of injured RGCs enhance neuronal survival and make possible a lengthy axonal regeneration that restores functional connections with the superior colliculus. These effects suggest that injured nerve cells in the mature central nervous system (CNS) are strongly influenced by interactions with components of their immediate environment as well as their targets. Under these conditions, injured CNS neurons can express capacities for growth and differentiation that resemble those of normally developing neurons. An understanding of this regeneration in the context of the cellular and molecular events that influence the interactions of axonal growth cones with their non-neuronal substrates and neuronal targets should help in the further elucidation of the capacities of neuronal systems to recover from injury.

Animals↗
PubMed1991

Construction and characterization of a partial library of yeast artificial chromosomes from human chromosome 21.

We report a protocol for cloning large DNA fragments in yeast artificial chromosomes (YAC). A partial library has been constructed from a somatic hybrid containing chromosome 21 as the single source of human DNA. About 4.0 Mb of human DNA was recovered in 17 YAC clones. Three clones were analyzed by in situ hybridization and mapped on chromosome 21. One clone hybridized with the chromosome 21 centromeric region and may provide new insight both on the molecular structure of centromere and on the localization of Alzheimer disease gene.

Base Sequence↗
PubMed1991

Neuronal and nonneuronal influences on retinal ganglion cell survival, axonal regrowth, and connectivity after axotomy.

In contrast to the abortive regrowth that occurs when axons are interrupted in the adult mammalian CNS, exposure of injured CNS axons to the nonneuronal milieu of a peripheral nerve can lead to extensive axonal elongation. With the application of this experimental approach to the retinocollicular pathway in adult rodents, it has been possible to investigate the influences of neuron-glia and other interactions on the capacity of axotomized CNS neurons to survive injury, to elongate the distances necessary to reach specific targets, and to form connections in the CNS in adult rodents. The results of these investigations indicate that the changed glial environment provided by peripheral nerve grafts permits the guided regeneration of RGC axons to their CNS targets. Back in the CNS glial environment, regenerated axons penetrate their targets for short distances and re-form normal appearing synapses that can excite or inhibit postsynaptic neurons. Further studies will require a better understanding of intrinsic neuronal properties and of the interactions of these neurons with other neurons and with the cellular and noncellular components of the extraneural milieu.

Animals↗
PubMed1991

Rapid changes in poly (A) tail length of vasopressin and oxytocin mRNAs form a common early component of neurohypophyseal peptide gene activation following physiological stimulation.

The time course of acute changes in vasopressin (VP) and oxytocin (OT) mRNA size and level during dehydration has been studied in rats. Total RNA was extracted from samples of the supraoptic nucleus at various intervals after water deprivation, subjected to northern blotting, and probed with oligonucleotides specific for VP and OT mRNA. The VP and OT mRNA size, shown previously to reflect 3'-poly (A) tail length, was consistently increased 2 h after dehydration, prior to significant changes in plasma osmolality or haematocrit. Intraperitoneal administration of hypertonic saline resulted in a similarly rapid VP and OT mRNA size response, in some cases within 1 h of treatment. The effect of a discrete hypovolaemic stimulus was investigated with intraperitoneal injections of polyethylene glycol; again, the VP and OT mRNA size was rapidly increased. No significant changes in mRNA level were observed in any of the experimental groups. The results show that an increase in VP and OT mRNA poly(A) tail length forms an acute and general response to activation of the hypothalamo-neurohypophyseal system. The rapidity of the poly (A) tail response, which appears to be independent of physiological signalling mechanisms associated with increases in mRNA accumulation (observed after 2 days of dehydration), provides a paradigm for the investigation of novel modes of neuronal gene regulation.

Animals↗
PubMed1991

The inner dynein arm complex: compatible images from freeze-etch and thin section methods of microscopy.

A study has been made of the inner dynein arm complex in the sperm flagellum of Gallus domesticus. It has been found that images of the complex made from highly contrasted, approx. 20 nm, sections are very largely compatible with images made from replicas of rapidly cryofixed material. This suggests that neither technique seriously distorts the native state. From both types of image, we conclude that the most proximal group of inner dynein arm heads (IDA 1) is related to spoke S1 and consists of 3 heads with fine connections to the B-tubule. IDA 2 consists of 2 such heads and is related to spoke S2. IDA 3 is apparently single headed and lies close to the A-attachment of spoke S3. This arrangement of IDAs repeats every 96 nm. Between the IDAs and outer dynein arms (ODAs), lying close to IDAs 2 and 3, is a strap-like linkage to the next B-tubule; it is argued that this represents the circumferential link, nexin. In sections, but not consistently in replicas, IDA2 lies closer to the plane of the nexin link than does IDA 1. In the presence of ATP and vanadate, little change is seen in the IDA complex except for an ill-defined alteration to IDA 1. It is speculated that the apparently smaller size of the inner arm heads (compared with the ODA major subunit) may have functional implications in relation to maximal sliding velocity.

Adenosine Triphosphate↗
PubMed1990

Vasopressin mRNA in parvocellular neurons of the rat suprachiasmatic nucleus exhibits increased poly (A) tail length following water deprivation.

Dehydration is associated with altered vasopressin (VP) gene expression in the rat hypothalamus; an increase in both arginine-vasopressin mRNA abundance and size (due to 3' poly(A) tail extension) has been observed previously. We have now shown that the effects of dehydration are not restricted to the magnocellular AVP neuronal systems since VP mRNA in the suprachiasmatic nucleus (SCN) exhibits a progressive increase in poly(A) tail length during dehydration. No significant increase in VP mRNA abundance was found. Similar kinetics of poly(A) tract extension in the SCN and magnocellular supraoptic nucleus implies a common regulatory mechanism which appears to be distinct from osmotic upregulation of VP transcript abundance.

Animals↗
PubMed1990

Temporally defined induction of c-fos in the rat pineal.

In the rat pineal gland c-fos is transiently and markedly induced after the onset of darkness. The induction is abolished following removal of the superior cervical ganglion and reproduced in ganglionectomized animals by treatment with the adrenergic agonist isoproterenol. These findings show for the first time that induction of c-fos forms an integral part of physiological changes in cellular activity. Expression of the c-fos gene may be functionally linked to the nocturnal changes in pineal indole metabolism.

Animals↗
PubMed1990

Regulation of c-fos and c-jun expression in the rat supraoptic nucleus.

1. We have investigated induction of the nuclear proto-oncogenes c-fos and c-jun in the rat supraoptic nucleus (SON) during physiological stimulation. 2. Dehydration (0-24 hr) was associated with modest, but significant increases in both c-fos and c-jun mRNA at 8 hr and 16 hr as determined by Northern analysis of total RNA extracted from microdissected SON. Prior to 8 hr, and beyond 24 hr, no consistent changes in c-fos and c-jun mRNA were found. Levels of c-fos and c-jun mRNA in the hippocampus were not altered over 24 hr of dehydration. 3. Acute stimulation with hypertonic saline (1.5 M, i.p.) resulted in a marked increase in SON c-fos mRNA at 1 hr (6-fold) and 2 hr (3.5-fold). Small increases in SON c-jun mRNA were observed at these time points. Treatment with a similar volume of 0.9% saline did not elevate SON c-fos and c-jun mRNA levels. 4. Analysis of transcriptional activity with a nuclear run-on assay showed that activation of transcription appears to mediate the induction of c-fos and c-jun mRNA following acute hypertonic saline treatment. During dehydration transcriptional activation is apparent for c-jun but is not well defined for c-fos. 5. The results are discussed with reference to the hypothesis that products of c-fos and c-jun may mediate adaptive changes in hypothalamic gene expression.

Animals↗
PubMed1990

Regrowth and connectivity of injured central nervous system axons in adult rodents.

The capacity of injured nerve cells to regrow and form terminal connections in the CNS of adult mammals was investigated in axotomized retinal ganglion cells (RGCs) of rodents whose optic nerves were substituted by an autologous segment of peripheral nerve. While many RGCs died after axotomy approximately 20% of the surviving RGCs regenerated axons several cm in length. Some of the regenerated RGC axons entered the superior colliculus where they arborized and formed well differentiated synapses that transynaptically excited or inhibited tectal neurons.

Animals↗