The cDNA for the Syrian hamster al-adrenergic receptor has been cloned with oligonucleotides corresponding to the partial amino acid sequence of the receptor protein purified from DDTMF-2 smooth muscle cells. The deduced amino acid sequence encodes a 515-residue polypeptide that shows the most sequence identity with the other adrenergic receptors and the putative protein product of the related clone G-21. Similarities with the muscarinic cholinergic receptors are also evident. Expression studies in COS-7 cells confirm that we have cloned the a!1-adrenergic receptor that couples to inositol phospholipid metabolism. a1-Adrenergic receptors have been implicated in a variety of functions, including smooth muscle contraction (1), regulation of hepatic glycogen metabolism (5, 6), and mitogenesis in certain tissues (7, 8).Our laboratory has cloned the genes and/or cDNAs for both subtypes of,-adrenergic receptor, 81 and /2(9-11), and for two different subtypes of the a2-adrenergic receptor (12, 13). Here we report the cloning and sequencinglI ofthe cDNA for the hamster a1-adrenergic receptor and its expression. This study brings to completion the cloning of the major subtypes of the adrenergic receptor family. METHODSPurification of the ar-Adrenergic Receptor. DDT1MF-2 cells were grown in 10 liters of suspension cultures as described (14). Cell membranes were prepared from about 1600 liters of cultured cells in 100-liter batches. From these membranes about 1 nmol of a1-adrenergic receptor was purified by affinity chromatography, wheat-germ agarose chromatography, and gel-permeation HPLC as reported (15).Peptide Purification and Amino Acid Sequencing. Purified a1-adrenergic receptor (1 nmol) was treated with 0.5 M CNBr/70% formic acid for 24 hr at 25°C. The digest was fractionated by reverse-phase HPLC with a Phenomenex (5 ,um, W-Porex) C4 column (4.6 x 250 mm) with a 10-80% acetonitrile gradient in 0.1% trifluoroacetic acid at a flow rate of 1 ml/min. Amino acid sequence analysis was performed by using an Applied Biosystems (Foster City, CA) 470A gasphase sequencer in combination with an Applied Biosystems 120A phenylthiohydantoin analyzer.Genomic and cDNA Library Screening. A hamster genomic DNA library in AEMBL3A was constructed as described (10). A hamster cDNA library was prepared from DDT1MF-2 poly(A)+ RNA. cDNA larger than 1.7 kilobases (kb) was purified by preparative agarose electrophoresis, ligated to the vector AgtlO (Stratagene, La Jolla, CA), and packaged in vitro (Gigapack Plus, Stratagene). Standard recombinant DNA and microbiological procedures were used (16). The oligonucleotides synthesized on Applied Biosystems 380 B DNA synthesizer were purified on a 16% denaturing polyacrylamide gel and were labeled with [y-32P]ATP at the 5'-hydroxyl group by phage T4 polynucleotide kinase. DNA fragments used as hybridization probes were labeled with [a-32P]dATP by random priming. Duplicate filters were hybridized in 0.90 M NaCI/0.09 M sodium citrate, pH 7/0.2% polyvinylpyrrolidone/0.2% Ficoll/0.2% bovine ser...

Further advances in the treatment of LUTS associated with BPH may depend not only on receptor subtype selectivity, but also on other pharmacokinetic and pharmacodynamic factors.

Human vascular alpha(1)AR subtype distribution differs from animal models, varies with vessel bed, correlates with contraction in mammary artery, and is modulated by aging. These findings provide potential novel targets for therapeutic intervention in many clinical settings.

Benign prostatic hyperplasia (BPH) is a common cause of urinary outflow obstruction in aging men leading to lower urinary tract symptoms (LUTS). a1-Adrenoceptors (a1ARs) antagonists (blockers) have become a mainstay of LUTS treatment because they relax prostate smooth muscle and decrease urethral resistance, as well as relieving bladder LUTS symptoms. A review of key recent clinical trials suggests new insights into the role of specific a1AR subtypes in the treatment of LUTS.

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It is unclear whether the regulatory distinction between non-identifiable and identifiable information — information used to determine informed consent practices for the use of clinically derived samples for genetic research — is meaningful to patients. The objective of this study was to examine patients’ attitudes and preferences regarding use of anonymous and identifiable clinical samples for genetic research. Telephone interviews were conducted with 1,193 patients recruited from general medicine, thoracic surgery, or medical oncology clinics at five United States academic medical centers. Wanting to know about research being done was important to 72% of patients when samples would be anonymous and to 81% of patients when samples would be identifiable. Only 17% wanted to know about the identifiable scenario but not the anonymous scenario (i.e., following the regulatory distinction). Curiosity-based reasons were the most common (37%) among patients who wanted to know about anonymous samples. Of patients wanting to know about either scenario, approximately 57% would require researchers to seek permission, whereas 43% would be satisfied with notification only. Patients were more likely to support permission (versus notification) in the anonymous scenario if they had more education, were Black, less religious, in better health, more private, and less trusting of researchers. The sample, although not representative of the general population, does represent patients at academic medical centers whose clinical samples may be used for genetic research. Few patients expressed preferences consistent with the regulatory distinction between non-identifiable and identifiable information. Data from this study should cause policy-makers to question whether this distinction is useful in relation to research with previously collected clinically derived samples.

The physiological significance of multiple (5, 6). In addition, the third intracellular loop and proximal cytoplasmic tail of the (32AR contain consensus sequences for PKAmediated phosphorylation (5), with the third intracellular loop site appearing to be the more critical for cAMP dependent desensitization (8). Examination of the molecular structure of the three ,BAR subtypes (9-11) reveals a striking difference between the P2-and f3AR. In contrast to the 11 serines and threonines in the distal portion of the cytoplasmic tail of the (32AR, there are only 3 serines in the same region ofthe 33AR.In addition to this apparent paucity of potential BARK phosphorylation sites, the 83AR lacks consensus sequences for PKA-mediated phosphorylation. Receptor sequestration and down-regulation are two other processes, independent of receptor phosphorylation, that also occur during agonist-promoted desensitization of the f82AR. Sequestration is a rapid (seconds to minutes) movement (internalization) of receptors from the cell surface to an intracellular compartment. As discussed below, the A3AR lacks an apparently critical 10-amino acid sequence motif in the cytoplasmic tail which has been implicated as playing a role in P2AR sequestration (12). Finally, after long-term (hours) agonist exposure, the net cellular expression of P2AR becomes decreased (regardless of localization), a process termed down-regulation. Two (but not necessarily all) of the requirements for full agonist-promoted down-regulation of the P2AR are PKA phosphorylation sites (13) and tyrosine residues in the cytoplasmic tail (14). Again, the 3AR lacks both of these determinants.Thus the ,B3AR appears to lack a number of the molecular features for agonist-promoted regulation which have thus far been established with the P2AR. This raises the interesting Abbreviations: 13AR, 1-adrenergic receptor; PARK, BAR kinase; PKA, cAMP-dependent protein kinase (protein kinase A);