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H J Cooke

Publications and source records attributed to H J Cooke.

At least 109 records · Page 6Linked to original sources

Hypervariable ultra-long telomeres in mice.

Telomere structure and behaviour is less well understood in vertebrates than it is in ciliates and yeasts (reviewed in ref. 1). Like all other eukaryotic chromosomes, those of vertebrates terminate in an array of a short repeated sequence. In vertebrates this sequence is (TTAGGG)n, as shown by in situ hybridization. In humans, these terminal repeats are heterogeneous in length, averaging about 10 kilobases in blood cells. Here we report the structure and inheritance of the terminal repeats present at mouse telomeres. The (TTAGGG)n tracts are many times larger than those present at human telomeres. Because of their constancy in length through somatic cell divisions, they are resolved as multiple discrete restriction fragments of up to 150 kilobases. Strikingly, this banding pattern is highly polymorphic within populations of inbred mice, suggesting an unusually high mutation rate. Indeed, although the banding pattern is inherited in a largely mendelian fashion, (TTAGGG)n tracts of new size appear frequently in family studies.

Animals↗

A VNTR immediately adjacent to the human pseudoautosomal telomere.

The probe 29C1 detects a hypervariable locus 18kb from the telomere of the human X and Y chromosomes, in the pseudoautosomal region. Here we report that hypervariability of fragments containing this sequence in the human population arises by loss or gain of a 31 base pair GC rich repeat. Labelled 29C1 does not detect a DNA fingerprint at low stringency, though the consensus repeat sequence does show some similarity to previously reported minisatellites. Sequence within the repeat block has G and C rich strands, a feature associated with sequences at the telomeres of many higher organisms. The repeat block shows sequence characteristics normally associated with a low methylation island, though the locus is methylated and does not appear to be transcribed.

Animals↗

Effects of tetrodotoxin on chloride secretion in rabbit distal colon: tissue and cellular studies.

The effects of tetrodotoxin (TTX) were examined in muscle-stripped segments of rabbit distal colon and in cells of isolated colonic crypts. Electrical field stimulation (EFS) of the submucosa/mucosa evoked an increase in short-circuit current (ISC) that was due to an increase in chloride secretion. The EFS-evoked response was reduced 81% by 10(-7) M TTX and 30% by 5 X 10(-6) M atropine. Vasoactive intestinal peptide (VIP), carbachol, aminophylline, and 1,1-dimethyl-4-phenylpiperazinium increased ISC. Bumetanide reduced the responses to neural stimulation, aminophylline, and VIP. To determine whether TTX had direct effects on crypt epithelial cells, crypts were isolated and cells were impaled with microelectrodes. Mean resting potential (Vbl) was -67 +/- 1.1 mV (n = 63). VIP and aminophylline depolarized Vbl by 34 +/- 4.6 (n = 13) and 34 +/- 3.5 mV (n = 18), respectively. TTX had no significant effect on resting Vbl or on the responses to VIP or aminophylline. We conclude that stimulation of submucosal neurons in the rabbit distal colon evokes a TTX- and bumetanide-sensitive increase in net chloride secretion that is dependent on the release of acetylcholine and other secretory neurotransmitters. Electrophysiological studies rule out a direct effect of TTX on colonic crypt cells.

Amiloride↗

Histamine augments colonic secretion in guinea pig distal colon.

We tested the hypothesis that the role of histamine in the control of intestinal secretion is mediated by prostaglandins (PGs). The effects of histamine on ion transport were examined in muscle-stripped sheets of mucosa/submucosa set up in flux chambers. Histamine evoked a transient concentration-dependent increase in short-circuit current (Isc) that was reduced by the Cl- transport inhibitor bumetanide. Histamine also caused the release of PGE2. The Isc response to histamine was reduced by indomethacin and piroxicam, which block PG formation, but not by nordihydroguaiaretic acid, which prevents production of lipoxygenase products. 2-Methylhistamine, but not dimaprit, evoked a concentration-dependent increase in Isc. The Isc response to histamine was reduced by the H1-blocker pyrilamine, but not by the H2-antagonist cimetidine. In addition to its direct effect, histamine augmented the responses of endogenously released neurotransmitters with and without indomethacin and hexamethonium. Tetrodotoxin (TTX) reduced the Isc response to 10(-3) M histamine. In the presence of TTX, exogenous histamine amplified the responses to PGs, vasoactive intestinal polypeptide, 2-chloroadenosine, bethanechol, and carbachol. These results suggest that histamine acts at H1-receptors on cells within the gut to mediate intestinal Cl- secretion in part by releasing PGs and by augmenting the actions of endogenously released neurotransmitters. Our results indicate that histamine has a role in the regulation of colonic transport function.

2-Chloroadenosine↗

H2 receptors mediate cyclical chloride secretion in guinea pig distal colon.

We tested the hypothesis that histamine mediates ion secretion in the guinea pig distal colon by stimulating H2 receptors on submucosal neurons. Serosal addition of histamine evoked a transient increase in short-circuit current (Isc) followed by recurrent cyclical increases in Isc. The transient phase of the response was examined previously and was not investigated in these studies. Histamine (1.5-2.5 x 10(-5) M) evoked a peak increase in Isc of 177 +/- 25 microA/cm2 at intervals of 5 min for 1-2 h. The duration of each recurrent cycle averaged 2.1 +/- 0.3 min. The H2 agonist dimaprit evoked recurrent cycles that had larger amplitudes than those caused by histamine. In the presence of histamine or dimaprit, the amplitude of the first cycle of the response was always less than subsequent cycles, regardless of the initial concentration. The cyclical responses to histamine, 2-methylhistamine, or dimaprit were unaltered by the H1 blocker pyrilamine, were reduced by the H2 antagonist cimetidine, and were abolished by the neuronal blocker tetrodotoxin. Blockade of prostaglandin formation with piroxicam did not prevent the recurrent cycles. The recurrent cycles were inhibited by the chloride transport blocker bumetanide and by removal of chloride ions. Our results demonstrate that histamine mediates prolonged cyclical chloride secretion in the guinea pig distal colon by activating H2 receptors on submucosal neurons involved in regulation of epithelial chloride transport.

Animals↗

Tachykinin-induced anion secretion in guinea pig distal colon: role of neural and inflammatory mediators.

The effects of substance P and related tachykinins on short-circuit current were determined in submucosa-mucosa sheets of guinea pig distal colon. Substance P, neurokinin A, eledoisin and physalaemin evoked a concentration-dependent increase in short-circuit current with a rank order of potencies of substance P = physalaemin greater than eledoisin = neurokinin A. The substance P response was reduced in chloride-free solutions or after addition of furosemide. Tetrodotoxin, atropine, mecamylamine and pyrilamine reduced the response to substance P. The substance P antagonist, [D-Arg1,D-Phe5,Trp7,9,Leu11]-substance P, reduced the response to neural stimulation and to exogenous substance P. The results suggest that substance P evokes anion secretion by activating cholinergic and noncholinergic neurons and by releasing histamine.

Animals↗

Cloning of human telomeres by complementation in yeast.

Telomeres confer stability on chromosomes by protecting them from degradation and recombination and by allowing complete replication of the end. They are genetically important as they define the ends of the linkage map. Telomeres of lower eukaryotes contain short repeats consisting of a G-rich and a C-rich strand, the G-rich strand running 5'-3' towards the telomere and extending at the end. Telomeres of human chromosomes share characteristics with those of lower eukaryotes including sequence similarity as detected by cross-hybridization. Telomeric repeats from many organisms can provide telomere function in yeast. Here we describe a modified yeast artificial chromosome (YAC) vector with only one telomere which we used to clone human telomeres by complementation in yeast. YACs containing human telomeres were identified by hydridization to an oligonucleotide of the trypanosome telomeric repeat. A subcloned human fragment from one such YAC is immediately subtelomeric on at least one human chromosome.

Animals↗

Thyrotropin-releasing hormone (TRH) evokes chloride secretion via cholinergic pathways in the guinea-pig ileum.

The effect of thyrotropin-releasing hormone (TRH) on ion transport in the guinea-pig ileum was investigated in flux chambers. TRH evoked a concentration-dependent increase in short-circuit current that was due to active anion secretion. The response to TRH was prevented by tetrodotoxin and nearly abolished by atropine, but was not affected by mecamylamine. The results suggest that TRH plays a role as a neuromodulator or neurotransmitter that influences secretory processes by directly or indirectly activating cholinergic motor neurons.

Animals↗

A search for restriction fragment length polymorphism on the human Y chromosome.

A systematic search for restriction fragment length polymorphisms (RFLPs) on the human Y chromosome was performed. DNA samples from 16-34 individuals were screened with five restriction enzymes and 12 Y-chromosomal probes, 3 of which detect lowly repetitive sequences and 9 of which are apparently single copy in genomic DNA. None of the single-copy probes revealed any variation. The repetitive sequence probe p21A1 (DYZ?) revealed a TaqI RFLP with q = 0.05. The frequency of fixed point mutations in Y-chromosomal DNA outside the pseudoautosomal region is probably less than 1 in 18,000 bp.

Blotting, Southern↗

Intestinal transport in megacolonic mice. Alterations in sugar absorption.

This study examined jejunal sugar absorption in piebald mice with congenital megacolon and in normal littermates. Transmural potential difference, short-circuit current, and tissue conductance of flat sheets of jejunum set up in flux chambers were significantly greater in the diseased mice compared to normal siblings. In piebald mice, net absorption of 3-o-methylglucose was enhanced due to a significant increase in mucosal-to-serosal flux compared with normal littermates. Stimulation of electrogenic sodium absorption by alanine (10 mM) increased basal short-circuit currents more in piebald tissues than in tissues from normal mice, whereas stimulation of ion transport by carbachol (10 microM) evoked an increase in short-circuit current that was similar in the two groups. Alterations in intestinal mass, morphology, or Na+, K+-ATPase activity could not account for the increase in absorptive function characteristic of piebald mice.

3-O-Methylglucose↗

Effects of enteric neural stimulation on chloride transport in human left colon in vitro.

Nonpathological segments of muscle-stripped left colon from patients undergoing surgery for carcinoma, diverticulitis, or other causes were set up in flux chambers in order to determine the influence of intrinsic neurons on ion transport. In the basal state, both sodium and chloride were actively absorbed, and there was no significant residual ion flux. Electrical field stimulation of the intrinsic mucosal innervation evoked an increase in short-circuit current that was dependent on stimulus frequency and strength. The response was mimicked by scorpion venom, which is known to depolarize neurons, and was nearly abolished by tetrodotoxin, which prevents neurotransmitter release. Atropine reduced, but did not abolish, the response to neural stimulation. Carbachol, aminophylline, and theophylline significantly increased short-circuit current above basal levels. Electrical field stimulation evoked an increase in short-circuit current that could be accounted for by a decrease in net chloride absorption without any significant effect on sodium absorption or residual ion flux. These results suggest that ion transport in the human left colon is regulated by intrinsic submucosal cholinergic neurons as well as other neuronal types, and activation of these nerves could provide the basis for certain diarrheal disorders.

Aged↗

Tonic activity of submucosal neurons influences basal ion transport.

The influence of tonically active submucosal neurons on basal ion transport was studied using sheets of guinea pig ileum set up in flux chambers. Tetrodotoxin evoked an immediate and sustained decrease in short-circuit current that was sustained for 60 minutes compared with control tissues in which basal currents gradually decreased over time. Time-dependent changes in basal short-circuit currents in tissues treated with atropine were not significantly different from control tissues. The decrease in short-circuit current after tetrodotoxin resulted from a greater increase in net chloride absorption than sodium absorption. Changes in net sodium and chloride transport were due to an increase in the mucosal-to-serosal fluxes of these ions. The results suggest that tonic activity of submucosal neurons limits the absorptive capacity of the guinea pig ileum.

Animals↗

Human alpha-calcitonin gene-related peptide influences colonic secretion by acting on myenteric neurons.

The effect of human alpha-calcitonin gene-related peptide (CGRP) on epithelial ion transport was investigated in guinea pig distal colon set up in Ussing flux chambers. Addition of CGRP to the serosal bathing solution evoked a dose-dependent increase in short-circuit current in whole-thickness tissues with intact myenteric and submucosal ganglia, but not in whole-thickness preparations when neural connections between myenteric and submucosal ganglia were severed, nor in sheets of submucosa/mucosa with intact submucosal ganglia. The effects of CGRP were nearly abolished in chloride-free solutions or after treatment with furosemide. Tetrodotoxin and hexamethonium abolished the effects of CGRP on basal short-circuit current whereas atropine did not. CGRP enhanced neurally evoked chloride secretion both in whole thickness and submucosa/mucosa preparations, but the effect in the latter was considerably smaller. These observations suggest that CGRP stimulates chloride secretion primarily by activating myenteric neurons that project either to submucosal ganglia or to the mucosa of the guinea pig distal colon. Furthermore, CGRP appears to have a greater effect on excitability of myenteric neurons than submucosal neurons.

Animals↗

Role of the "little brain" in the gut in water and electrolyte homeostasis.

The enteric nervous system plays a key role in maintenance of body fluid homeostasis by regulating the transport of ions by the intestinal epithelium. The epithelial cells normally absorb large volumes of fluid and ions daily, but tonically active submucosal neurons continuously suppress ion transport and limit the absorptive capacity of the intestine. Specialized nerve endings detect chemical, osmotic, or thermal alterations of the luminal contents or mechanical activity of the gut wall and encode this information as action potentials that propagate along nerve processes to the ganglia. Information transfer within the ganglia occurs at nicotinic cholinergic or other synapses. Ion transport is altered when neurotransmitters released from motor neurons interact with receptors on epithelial cells to initiate stimulus-response coupling. The signals that transduce changes in epithelial ion transport are largely unknown, except for acetylcholine, but may include vasoactive intestinal peptide or other peptides. These trigger changes in intracellular messengers that influence the state of ionic channels in the epithelial cells and thereby inhibit absorptive processes or stimulate secretory mechanisms. When conservation of salt and water is necessary, command signals from the central nervous system, and perhaps from the myenteric ganglia, will shut down the synaptic circuits in the submucosal ganglia and enhance the absorptive capacity of the bowel.

Animals↗

Neuromodulation of intestinal transport in the suckling mouse.

The influence of enteric nerves on intestinal ion and sugar transport was investigated in 3- to 4-wk-old suckling mice. Whole thickness segments of jejunum were mounted as flat sheets in flux chambers equipped to electrically stimulate nerves in the intestinal wall. Tetrodotoxin significantly reduced basal short-circuit current in a subset of tissues containing 3-O-methylglucose in the mucosal bath. Electrical field stimulation of intrinsic nerves evoked an increase in short-circuit current of 86 +/- 15 microA/cm2 that was due to an increase in active chloride secretion with no significant change in the net movement of other ions. The secretory response to neural stimulation was abolished by tetrodotoxin and reduced by the muscarinic antagonist, atropine. Mucosal-to-serosal fluxes of 3-O-methylglucose were not altered by the cholinergic agonist carbachol. These results show that ion transport in the suckling mouse jejunum is regulated by the enteric nervous system. Neural stimulation evokes a chloride secretory response that is mediated by acetylcholine and other noncholinergic transmitters. Sugar transport in the murine small intestine does not appear to be influenced by muscarinic cholinergic agonists.

3-O-Methylglucose↗

A human Y-chromosomal DNA sequence expressed in testicular tissue.

Clone pJA36B (DYS14) was isolated from a human Y chromosome enriched cosmid library. Southern blot analysis revealed a male-specific hybridization pattern. Deletion mapping with patients' DNA localized pJA36B to the median region of Yp, being present in the DNA of nine of fifteen XX-males tested so far and therefore localized in the region neighbouring the TDF-locus. Northern blot analysis showed a transcription signal in poly(A)+ RNA of human testis. Sequence analysis of the genomic DNA sequence revealed an open reading frame of 522 basepairs in the absence of control or signal sequences for the regulation of transcription or polyadenylation. This suggests that only one exon of a translatable sequence is present in clone pJA36B. A computer aided search revealed no significant homologies with known DNA or protein sequences.

Amino Acid Sequence↗

Evolution of homologous sequences on the human X and Y chromosomes, outside of the meiotic pairing segment.

A sequence isolated from the long arm of the human Y chromosome detects a highly homologous locus on the X. This homology extends over at least 50 kb of DNA and is postulated to be the result of a transposition event between the X and Y chromosomes during recent human evolution, since homologous sequences are shown to be present on the X chromosome alone in the chimpanzee and gorilla.

Animals↗