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Biomedical subjects

V Venkataraman

Publications and source records attributed to V Venkataraman.

35 records · Page 2Linked to original sources
PubMed1998

The alpha(2D/A)-adrenergic receptor-linked membrane guanylate cyclase: a new signal transduction system in the pineal gland.

In the pineal gland, the membrane guanylate cyclase activity was specifically stimulated by alpha(2D/A)-adrenergic receptor (alpha(2D/A)-AR) agonists. The agonists, however, did not stimulate the cyclase activity in the cell-free membranes. It was possible to stimulate the cyclase in cell-free membranes by the addition of the pineal soluble fraction, but this stimulation was Ca2+-dependent and alpha(2D/A)-agonist-independent. It was also possible to achieve Ca2+-dependent stimulation of the cyclase by the direct addition of CD-GCAP to the isolated pineal membranes. CD-GCAP is a Ca2+-binding protein and is a specific activator of one of the two members of the ROS-GC subfamily of membrane guanylate cyclases, ROS-GC1. The soluble fraction of the pineal gland stimulated recombinant ROS-GC1 in a Ca2+-dependent fashion. The direct presence of both ROS-GC1 and CD-GCAP in the pineal was established by molecular cloning/PCR studies. The findings demonstrate the existence of a novel signal transduction mechanism--the linkage of the alpha(2D/A)-AR signaling system with ROS-GC1 transduction system, occurring through intracellular Ca2+ via CD-GCAP.

Animals↗
PubMed1998

Rod outer segment membrane guanylate cyclase type 1 (ROS-GC1) gene: structure, organization and regulation by phorbol ester, a protein kinase C activator.

At present there are two recognized members of the ROS-GC subfamily of membrane guanylate cyclases. They are ROS-GC1 and ROS-GC2. A distinctive feature of this family is that its members are not switched on by the extracellular peptide hormones; instead, they are modulated by intracellular Ca2+ signals, consistent to their linkage with phototransduction. An intriguing feature of ROS-GC1, which distinguishes it from ROS-GC2, is that it has two Ca2+ switches. One switch inhibits the enzyme at micromolar concentrations of Ca2+, as in phototransduction; the other, stimulates. The stimulatory switch, most likely, is linked to retinal synaptic activity. Thus, ROS-GC1 is linked to both phototransduction and the synaptic activity. The present study describes (1) the almost complete structural identity of 18.5 kb ROS-GC1 gene; (2) its structural organization: the gene is composed of 20 exons and 19 introns with classical GT/AG boundaries; (3) the activity of the ROS-GC1 promoter assayed through luciferase reporter in COS cells; and (4) induction of the gene by phorbol ester, a protein kinase C (PKC) activator. The co-presence of PKC and ROS-GC1 in photoreceptors suggests that regulation of the ROS-GC1 gene by PKC might be a physiologically relevant phenomenon.

Animals↗
PubMed1997

The bovine alpha 2D-adrenergic receptor gene: structure, expression in retina, and pharmacological characterization of the encoded receptor.

This report describes cloning of the bovine alpha 2D-adrenergic receptor (alpha 2D-AR) gene and determination of the transcription start site, unequivocal presence of the alpha 2D-AR transcript in the retina, and pharmacological characteristics of the encoded product. Furthermore, expression of the gene in selected bovine tissues has also been scrutinized. A genomic clone was isolated from lambda EMBL3 library and a 3 kb fragment was subcloned and sequenced. This fragment contained the putative TATA box and the coding region. The encoded receptor was transiently expressed in COS cells. The recombinant receptor expressed pharmacological characteristics almost identical to the wild-type bovine retinal receptor, which were typical of the alpha 2D-AR subtype. RNase protection analysis confirmed the expression of the gene in the retina. The bovine receptor was structurally close to its rat analogue which also encodes the alpha 2D-AR, but, the highest homology was observed with the porcine receptor expressing alpha 2A-AR pharmacological characteristics. Certain structural features of the bovine gene were unique to itself and not shared by any other alpha2-AR subtype. Among the tissues tested using reverse transcriptase-polymerase chain reaction (RT-PCR), the alpha 2D-AR message was the most abundant in retina, followed by the brain and olfactory lobe. Thus, the availability of the bovine receptor gene probe will become an important additional tool in the elucidation of molecular mechanisms behind the alpha 2D-AR physiology in neurosensory processes such as those occurring in the eye and the brain.

Amino Acid Sequence↗
PubMed1997

A role for amino acid residues in the third cytoplasmic loop in defining the ligand binding characteristics of the alpha2D-adrenergic receptor.

In this report, 5 amino acid residues (aa) in the third cytoplasmic loop of the alpha 2D-adrenergic receptor are identified which (individually or together) alter its ligand-binding characteristics. An important structural discrepancy exists in the third cytoplasmic loop of the alpha 2D-ARs encoded by the rat cDNA and the rat gene--five aa are different. The newly identified bovine receptor as well as the mouse receptor contained the 5 aa identical to that encoded by the rat cDNA. Site-directed mutation of these residues to those of the rat gene encoded receptor resulted in alteration of binding characteristics: significant changes in the ability of the mutant receptor to bind to a number of agonists and antagonists were observed--ranging from a decrease by half in the case of oxymetazoline, to near total loss of binding in the case of prazosin. Thus, the mutant receptor was no longer pure alpha 2D-AR. This indicated a hitherto unrealized role of the third cytoplasmic loop in defining the ligand-binding characteristics of the receptor, and also suggested that the rat gene sequence was most probably in error.

Amino Acid Sequence↗
PubMed1997

Safety pharmacology of a combination of tinidazole and oxyphenonium bromide.

The effect of a combination of 150 mg tinidazole (CAS 19387-91-8) and 1 mg oxyphenonium bromide (CAS 50-10-2), referred to as the combination, was examined in various animal species to assess its safety. In mice and rats, the combination in the dose range 30-1000 mg/kg p.o. or 250 mg/kg i.p. did not produce behavioural or neurological changes, nor did it influence pentobarbital sleeping time, though, alcohol sleeping time was altered. In anaesthetised dogs, neither blood pressure, ECG, heart rate, respiration or gastrointestinal motility was affected after single intraduodenal administration of the combination 20 and 50 mg/kg or after chronic oral administration of 25 mg/kg daily for 15 days. In isolated organs, viz. perfused rabbit heart, guinea-pig ileum and rat ileum no significant changes were observed following various doses of the combination, compared to tinidazole and oxyphenonium bromide given alone in corresponding concentrations. In conclusion, the pharmacological profile of the aforementioned combination in the dosages employed in mice, rats, guinea-pigs, rabbits and dogs shows that it is safe and well tolerated.

Animals↗
PubMed1996

Molecular and pharmacological identity of the alpha 2D-adrenergic receptor subtype in bovine retina and its photoreceptors.

The rat cA2-47 gene encodes the pharmacologically defined alpha 2D-adrenergic receptor (alpha 2D-AR) subtype. Previously, the expression of its mRNA was shown in bovine retina by amplification through the reverse transcription-polymerase chain reaction (RT-PCR) of a region corresponding to the rat alpha 2D-AR, amino acid (aa) residues 382-439, indicating the presence of this subtype in this neural tissue. In the present study, the structure of this gene has been probed and the encoded receptor subtype has been characterized in bovine retina and its photoreceptor cells. The deduced aa sequence of the two bovine gene fragments, aa residues 290-375 and aa residues 392-434, demonstrates 77% overall identity with the rat alpha 2D-AR subtype and 80% overall identity with the mouse alpha 2D-AR. The receptor encoded by the bovine gene was expressed in the retina and its photoreceptors with the typical pharmacological characteristics established for the rat alpha 2D-AR subtype: The receptor bound rauwolscine with a KD of 14 nM in the retina and with that of 19 nM in the photoreceptor cells; the binding association rate constant, k+1, for the ligand was 0.012 min-1, the dissociation rate constant, k-1, was 0.14 min-1 and the half-time for dissociation was 5 min. Oxymetazoline displaced the bound [3H]-rauwolscine with an EC50 value of 85 nM, while SK & F 104078, and prazosin displaced the bound [3H]-rauwolscine with the respective IC50 values of 900 nM and 3000 nM. The other alpha 2-AR subtypes -alpha 2A-AR, alpha 2B-AR, alpha 2C-AR-were not detected in the retina and its photoreceptors. Thus, this study shows that the bovine alpha 2D-AR gene is a structural variant of the rat and mouse genes, that the bovine gene encodes the typical pharmacologically defined alpha 2D-AR subtype, that this subtype is present in its exclusive form in the bovine retina and its photoreceptors, where it may be presynaptic in nature.

Adrenergic alpha-Agonists↗
PubMed1996

A comparison of phase encoding ordering schemes in T2-weighted GRASE imaging.

Gradient and spin echo (GRASE) imaging is an echo train imaging sequence that combines gradient and RF refocusing. This combination introduces phase modulations into the echo train. If the phase encoding order is linear with echo time, these modulations cause severe ghosting artifacts. Changing the order of phase encoding can greatly reduce these artifacts. Several phase encoding orders for T2-weighted sequences are compared in this paper, linear, partially randomized, standard GRASE ordering, and k-banded (kb) GRASE ordering. Different possible implementations of GRASE and kbGRASE are also considered. Computer simulation is used to compare resolution and artifact levels. Phantom and volunteer images are presented. The linear order is most sensitive to ghosting artifacts associated with chemical shift, susceptibility differences and static field inhomogeneities. The standard GRASE order is least sensitive to these but most vulnerable to artifacts associated with short T2 signals, kb-GRASE is a good intermediate between linear and standard GRASE and generally shows the lowest artifact levels. The partially randomized order gives the most diffuse artifacts. Computer simulations show that spatial resolution and contrast with all phase encoding orders are similar.

Artifacts↗
PubMed1994

Structure, expression and gene sequence of a juvenile hormone esterase-related protein from metamorphosing larvae of Trichoplusia ni.

A carboxylesterase with an encoded molecular size of 61 kDa and a high sequence similarity to juvenile hormone esterase (JHE) has been cloned from cDNA prepared from final instar larvae of Trichoplusia ni. The absence of a recognizable encoded signal peptide suggests that the enzyme, JHER (for JHE-related) may not be secreted, in contrast to JHE. When the amino acid sequence of JHE, JHER and other esterases were mapped onto the secondary and tertiary structure determined crystallographically for acetylcholinesterase, certain structural features for the substrate binding/catalytic site were identified as common only to JHE and JHER. However, several differences between JHE and JHER were identified in residues at the binding/catalytic site, suggesting that although the two enzymes prefer similar natural substrates, these substrates are not identical. JHER is present as a single-copy gene, transcribed during the feeding stage of the final stage of the final larval stadium, but not after metamorphic commitment to the pupal developmental programme. The gene transcribes a single-size message of 2.0 kb. The genes for JHER and JHE appear to be physically juxtaposed in the T. ni genome. The 5' flanking sequence to the JHER gene possesses some sequences in common with the JHE gene, but is also missing some regulatory elements previously identified in the JHE gene. Sequences conserved between the promoters for the two genes were identified that were different from previously reported regulatory elements of eukaryotic transcription factors.

Acetylcholinesterase↗
PubMed1994

Regulation of juvenile hormone esterase gene transcription by juvenile hormone.

Juvenile hormone (JH) is a major hormone regulating insect development. We have obtained a cDNA and a genomic clone for juvenile hormone esterase (JHE), the enzyme that is involved in the degradation of juvenile hormone and which is critical for insect development. Analysis of the regulation of JHE during the final larval stadium in the cabbage looper, Trichoplusia ni, showed that the JHE mRNA levels are maximal on days 2 and 4 of the final stadium. Nuclear run-on analyses demonstrated that changes in JHE mRNA levels are primarily due to changes in the transcription rate of the gene, which may be a single copy in the genome. Treatment with a JH analog resulted in induction of JHE gene transcription, which could be detected within three hours after treatment. Salient features present in the 5' flanking region of this JH-sensitive gene are presented, including the presence of sequences closely resembling binding sites for members of the family of nuclear receptors. This report is the first direct demonstration, by nuclear run-on analysis, of JH induction of gene transcription.

Amino Acid Sequence↗
PubMed1993

Juvenile hormone action to suppress gene transcription and influence message stability.

Proteins normally expressed in high abundance only at larval-pupal metamorphosis in Trichoplusia ni were examined in a comparative analysis of the role and level of hormonal control of their expression. Some related proteins in the hemocyanin-superfamily (i.e., an acidic protein [AJHSP1] and two basic proteins [BJHSP1, BJHSP2]) were shown by nuclear run-on analysis to be specifically transcriptionally suppressed by juvenile hormone (JH), while transcription of another member of that family which is also metamorphosis-associated (arylphorin) was not specifically sensitive to JH. The stability of the mRNA for those members transcriptionally down-regulated by JH appeared to decrease under high JH conditions. While each protein was resorbed to some extent by the prepupal fat body, only the two basic proteins were quantitatively cleared from prepupal hemolymph. The JH-sensitive proteins studied appear to be encoded in single copy genes not immediately juxtaposed in the genome. These and previous studies now permit a more comprehensive understanding of the different combinations of mechanisms involving transcription, mRNA stability, translation, and protein clearance that operate to regulate these metamorphosis-associated proteins.

Animals↗
PubMed1993

Transcriptional regulation of an unusual trypsin-related protein expressed during insect metamorphosis.

The full-length cDNA for a trypsin-related protein expressed during larval-pupal metamorphosis was obtained. The encoded N-terminal half of the protein possessed similarity to trypsin proteases, including the correctly positioned histidine and aspartic acid members of the catalytic triad. However, the remainder of the encoded protein bore little resemblance to trypsin, and was altogether missing the canonical sequence containing the catalytic serine residue. The product of in vitro transcription/translation of the cDNA was a 28 kDa protein. Under normal conditions of a declining juvenile hormone titer, transcription of the mRNA as a proportion of total genomic transcription steadily increased for the first 3 days of the final stadium, but this increase was delayed and suppressed by maintenance of a high juvenile hormone titer. During the normal increase in transcription on day 3 of the final stadium, a sharp decline was observed in the steady-state abundance of the transcript, measured relative to abundance of the remaining mRNAs, suggesting that the stability of the message decreases after tissue commitment for a pupal molt.

Amino Acid Sequence↗
PubMedDate not supplied

Single-shot GRASE imaging with short effective TEs.

A new phase-encoding scheme for gradient- and spin-echo (GRASE) imaging giving a short effective TE is described. Unlike previous orders, phase encoding is centric rather than sequential. The sequence is a development of k-banded GRASE that uses different time segments of the echo train to encode different bands of k space. This phase-encoding order has been implemented in single-shot sequences on an imager with high performance gradients. Approximately 144 phase-encoding lines can be acquired in an echo train time of 390 ms. With centric phase encoding, the effective TE is 8 ms, compared with 75 ms for sequential encoding, and signal-to-noise ratios (SNRs) in brain tissue are 50 to 70% higher. The sequence can be employed in, for example, diffusion and velocity imaging.

Artifacts↗
PubMedDate not supplied

Adrenal cortical function and reserve in lepromatous leprosy.

22 chronic Lepromatous Leprosy patients of over 10 years duration, 17 non-reactional and 5 in reactional state who have not taken steroids as part of treatment were selected for the study. Serum cortisol was estimated by Radio-immuno-assay. Samples for basal values were collected at 8.00 A.M. Stimulated values were estimated in samples collected 8 hours after ACTH gel 40 units IM or 2 hours after 0.15 unit/kg BW Plain Insulin I.V. Basal cortisol values are: Normal controls 123.06 +/- 57.33 ng/ml. Non-reactional: 100.47 +/- 30.33 ng/ml; Reactional 141.4 +/- 43.15 ng/ml. Stimulated values are: Normal controls: 207.6 +/- 72.57 ng/ml. Non-reactional: 175.33 +/- 57.07 ng/ml, Reactional: 230 +/- 40.92 ng/ml. Basal serum cortisol in the non-reactional state is slightly lower than in normals but not statistically significant (P greater than 0.1). The basal cortisol in reactional subjects is slightly higher than in normals but not significant statistically (P greater than 0.05). The percentage rise over the basal value after stimulation test is found to be significantly low in both the reactional and non reactional states (P greater than 0.05) and also there is no statistically significant difference between these two groups (p greater than 0.5). Hence it is concluded that the Adrenal cortical reserve is low both in the non-reactional and reactional states of Lepromatous Leprosy.

Adrenal Cortex↗