In utero gene therapy rescues microcephaly caused by Pqbp1-hypofunction in neural stem progenitor cells.

Human mutations in PQBP1, a molecule involved in transcription and splicing, result in a reduced but architecturally normal brain. Examination of a conditional Pqbp1-knockout (cKO) mouse with microcephaly failed to reveal either abnormal centrosomes or mitotic spindles, increased neurogenesis from the neural stem progenitor cell (NSPC) pool or increased cell death in vivo. Instead, we observed an increase in the length of the cell cycle, particularly for the M phase in NSPCs. Corresponding to the developmental expression of Pqbp1, the stem cell pool in vivo was decreased at E10 and remained at a low level during neurogenesis (E15) in Pqbp1-cKO mice. The expression profiles of NSPCs derived from the cKO mouse revealed significant changes in gene groups that control the M phase, including anaphase-promoting complex genes, via aberrant transcription and RNA splicing. Exogenous Apc4, a hub protein in the network of affected genes, recovered the cell cycle, proliferation, and cell phenotypes of NSPCs caused by Pqbp1-cKO. These data reveal a mechanism of brain size control based on the simple reduction of the NSPC pool by cell cycle time elongation. Finally, we demonstrated that in utero gene therapy for Pqbp1-cKO mice by intraperitoneal injection of the PQBP1-AAV vector at E10 successfully rescued microcephaly with preserved cortical structures and improved behavioral abnormalities in Pqbp1-cKO mice, opening a new strategy for treating this intractable developmental disorder.

Design of JT-60SA core Thomson scattering diagnostic system.

An incoherent Thomson scattering diagnostic will be installed in the JT-60SA tokamak to measure electron temperature and electron density profiles. The target radial spatial resolution is 25 mm with 46 spatial channels. The accuracy in electron temperature and density is a few percent at ne = 7.5 × 1019 m-3, which is the expected value in the plasma core. This paper presents the designs of collection optics, fibers with their alignment system, and polychromators. The collection optics overcomes unique issues for superconducting fusion devices, i.e., limited design space, high-temperature measurements, and harsh radiation condition. When in several years the more performing plasma will generate intense nuclear radiation, the lens materials of the optics can be replaced by radiation resistant glasses without major changes in the lens holder. It will prevent transmission degradation and keep stable measurement accuracy.

Periodic metallo-dielectric structure in diamond.

Intense ultrashort light pulses induce three dimensional localized phase transformation of diamond. Photoinduced amorphous structures have electrical conducting properties of a maximum of 64 S/m based on a localized transition from sp(3) to sp(2) in diamond. The laser parameters of fluence and scanning speed affect the resultant electrical conductivities due to recrystallization and multi-filamentation phenomena. We demonstrate that the laser-processed diamond with the periodic cylinder arrays have the characteristic transmission properties in terahertz region, which are good agreement with theoretical calculations. The fabricated periodic structures act as metallo-dielectric photonic crystal.

Follow-up of a self-resolving renal pseudoaneurysm using contrast-enhanced power Doppler ultrasonography.

It has been standard practice to embolise a pseudoaneurysm caused by penetrating trauma whenever it is found, even in the absence of overt symptoms. This is a case report of a renal pseudoaneurysm (RPA) caused by a stab wound, which was safely monitored and followed using colour Doppler ultrasonography. On day 1, angiography showed a pseudoaneurysm of the renal artery in the parenchyma and ultrasonography showed blood flow into the pseudoaneurysm. Although abnormal blood flow into the kidney was seen, there appeared to be no leakage of blood from the pseudoaneurysm. The abnormal flow disappeared on day 12 and the area of the pseudoaneurysm became unclear from day 13. This report suggests the possibility that RPA caused by a stab wound could be an indication for conservative therapy under the following conditions: the RPA is detected initially; close monitoring using a colour Doppler ultrasound is possible; there is no leakage of blood from the right subclavian artery and there is a 2-week period of observation.

Hyperactive mutants of mouse D-aspartate oxidase: mutagenesis of the active site residue serine 308.

The role of Ser-308 of murine D-aspartate oxidase (mDASPO), particularly its side chain hydroxyl group, was investigated through the use of site-specific mutational analysis of Ser-308. Recombinant mDASPO carrying a substitution of Gly, Ala, or Tyr for Ser-308 was generated, and fused to either His (His-mDASPO), or glutathione S-transferase, His, and S (GHS-mDASPO) at its N-terminus. Wild-type His-mDASPO or GHS-mDASPO or their mutant derivatives were expressed in Escherichia coli and purified by affinity chromatography. All purified recombinant proteins had functional DASPO activity. The Gly-308 and Ala-308 mutants had significantly higher catalytic efficiency towards D-Asp and N-methyl-D-Asp, and a higher affinity for flavin adenine dinucleotide (FAD) compared to the wild-type enzyme. The Tyr-308 mutant had lower catalytic efficiency and binding capacity. These results suggest that the side chain hydroxyl group of a critical residue of mDASPO, Ser-308, down-regulates enzymatic activity, substrate binding, and FAD binding. This study provides information on the active site of DASPO that will considerably enhance our understanding of the biological significance of this enzyme.

The hnRNP-Htt axis regulates necrotic cell death induced by transcriptional repression through impaired RNA splicing.

In this study, we identify signaling network of necrotic cell death induced by transcriptional repression (TRIAD) by α-amanitin (AMA), the selective RNA polymerase II inhibitor, as a model of neurodegenerative cell death. We performed genetic screen of a knockdown (KD) fly library by measuring the ratio of transformation from pupa to larva (PL ratio) under TRIAD, and selected the cell death-promoting genes. Systems biology analysis of the positive genes mapped on protein-protein interaction databases predicted the signaling network of TRIAD and the core pathway including heterogeneous nuclear ribonucleoproteins (hnRNPs) and huntingtin (Htt). RNA sequencing revealed that AMA impaired transcription and RNA splicing of Htt, which is known as an endoplasmic reticulum (ER)-stabilizing molecule. The impairment in RNA splicing and PL ratio was rescued by overexpresion of hnRNP that had been also affected by transcriptional repression. Fly genetics with suppressor or expresser of Htt and hnRNP worsened or ameliorated the decreased PL ratio by AMA, respectively. Collectively, these results suggested involvement of RNA splicing and a regulatory role of the hnRNP-Htt axis in the process of the transcriptional repression-induced necrosis.

A model of plasma current through a hole of Rogowski probe including sheath effects.

In TST-2 Ohmic discharges, local current is measured using a Rogowski probe by changing the angle between the local magnetic field and the direction of the hole of the Rogowski probe. The angular dependence shows a peak when the direction of the hole is almost parallel to the local magnetic field. The obtained width of the peak was broader than that of the theoretical curve expected from the probe geometry. In order to explain this disagreement, we consider the effect of sheath in the vicinity of the Rogowski probe. A sheath model was constructed and electron orbits were numerically calculated. From the calculation, it was found that the electron orbit is affected by E × B drift due to the sheath electric field. Such orbit causes the broadening of the peak in the angular dependence and the dependence agrees with the experimental results. The dependence of the broadening on various plasma parameters was studied numerically and explained qualitatively by a simplified analytical model.

Large deletion of androsterone UDP-glucuronosyltransferase gene in the inherited deficient strain of Wistar rats.

LA Wistar rats have a deficiency of androsterone UDP-glucuronosyltransferase (UDPGT) and are present in Wistar rat colonies around the world. In order to clarify the molecular mechanism of the deficiency, androsterone UDPGT cDNA clone, pGT2 was isolated from rat liver cDNA library and was digested with restriction enzymes to afford three probes for Northern and Southern blot analyses in HA (normal), heterozygous LA and LA Wistar rats. In Northern blot analysis, androsterone UDPGT mRNA was totally absent in LA Wistar rat liver. Southern blot analysis suggested a large deletion of androsterone UDPGT gene in the rats. Genomic DNA amplifications with synthetic primers which have nucleotide sequences corresponding to the 5'-region of androsterone UDPGT cDNA, suggested that androsterone UDPGT gene has exon 1 with a length of some 700 bp and that this exon is deleted in LA Wistar rats. Based on these lines of evidence, it is concluded that the large portion of androsterone UDPGT gene is deleted in LA Wistar rats, which results in the absence of androsterone UDPGT mRNA and consequently the corresponding enzyme protein.

Immunochemical characterization of developmental changes in rat hepatic hydroxysteroid sulfotransferase.

A major isoenzyme of hepatic androsterone-sulfating sulfotransferase (AD-ST) was purified from adult female rats. The activity was purified 122-fold over that found in the cytosol and showed a single protein band with a subunit molecular mass of 30 kDa after sodium dodecyl sulfate polyacrylamide gel electrophoresis. The purified enzyme exhibited four isoelectric variants of subunits on denaturing isoelectrofocusing gels (pI = 5.8, 6.1, 6.7 and 7.2). Rabbit antiserum raised against the enzyme specifically detected AD-ST polypeptide in rat liver cytosol. Immunoblot analysis of liver cytosol from female and male rats at various ages showed good correlation between the levels of AD-ST activity and AD-ST polypeptide. Significant levels of AD-ST activity and polypeptide were detected in senescent male rats, though normal adult male rats have very low levels of AD-ST activity and protein. The relative content of the isoelectric variants of AD-ST were different in liver cytosol of weanling and adult females, indicating that age- and gender-related alterations of hepatic AD-ST activity are primarily determined by the levels of AD-ST polypeptide and the relative amounts of the four isoelectric variants of the enzyme.

Regional distribution and postnatal changes of D-amino acids in rat brain.

Regional distribution of D-amino acids in rat brain was studied by the modified highly sensitive analytical method which was previously developed. The method includes fluorogenic derivatization of each amino acid, isolation of each amino acid by reverse-phase HPLC, followed by enantiomeric separation with Pirkle-type chiral stationary phases. D-Amino acid contents were determined in the cerebrum, cerebellum, hippocampus, medulla oblongata, pituitary gland and pineal gland. D-Aspartic acid was observed in the pineal gland (3524 +/- 263 nmol/g, data are for male rats of 6 weeks of age) and the pituitary gland (80.5 +/- 9.0 nmol/g). D-Serine was found in various regions of the brain except for the cerebellum and medulla oblongata. D-Alanine was observed exclusively in the pituitary gland (25.9 +/- 4.4 nmol/g), whereas D-leucine was found in the pineal gland (3.4 +/- 0.4 nmol/g) and the hippocampus (1.6 +/- 0.07 nmol/g). No other D-amino acids were detected in the brain. The contents of D-aspartic acid in the pituitary gland and D-serine in the pineal gland were higher in female rats. In contrast the contents of D-alanine in the pituitary gland and D-leucine in the pineal gland and the hippocampus were higher in males. Postnatal changes of D-aspartic acid and D-leucine in the pineal gland and D-alanine in the pituitary gland were also investigated. The results described in this paper suggested that distinct regulatory mechanisms exist for individual D-amino acids in the corresponding region of rat brain.

Incorporation and metabolism of 2-acyl lysophospholipids by Escherichia coli.

The incorporation of 2-acyl lysophospholipids into Escherichia coli, and their metabolism were studied. 2-[14C]Acyl lysophosphatidylethanolamine could penetrate into E. coli cells and was mainly incorporated into phosphatidylethanolamine. 2-Acyl lysophosphatidylethanolamine was partially degraded, but some of it was incorporated into membrane phospholipids by acylation. 2-Acyl lysophosphatidylcholine also entered cells and was acylated to phosphatidylcholine. The acylation of 2-acyl lysophospholipid by the envelope fraction was also studied. Fatty acids were incorporated into 2-acyl lysophospholipids by the envelope fraction in the presence of ATP and Mg2+, and the incorporation was stimulated by acyl carrier protein, but not by coenzyme A. No acylation was observed with acyl coenzyme A as acyl donor. The acylation activities of the inner and outer membranes were examined. Pathways for degradation and modification of membrane phospholipids in E. coli are proposed.

Site-directed mutagenesis of rat hepatic hydroxysteroid sulfotransferases.

Two cDNA clones of rat hepatic hydroxysteroid sulfotransferase (ST) (ST-40 and ST-20) were isolated and expressed in Escherichia coli cells. Several histidine residues in their coding regions are highly conserved in the ST superfamily, and histidine mutants were constructed by site-directed mutagenesis. The substitution of alanine or lysine for the histidine at position 98 in the ST-40 enzyme resulted in a loss of ST activities toward dehydroepiandrosterone (DHEA), androsterone (AD) and cortisol (CS). The mutation of histidine 98 into alanine abolished the specific binding to 3'-phosphoadenosine 5'-phosphate agarose, suggesting that the residue is located at a critical position in the 3'-phosphoadenosine 5'-phosphosulfate (PAPS) binding site. In the ST-20 enzyme, the replacement of histidine 98 with alanine also resulted in the loss of ST activity toward its preferential substrate, CS. In the ST-40 enzyme, the mutation at histidine 256 into alanine markedly reduced CS-ST activity, but DHEA-ST activity was not changed. Furthermore, selective decrease in CS-ST activity was also observed in the alanine mutant at lysine 254 or at asparagine 255 of the ST-40 enzyme. Kinetic analysis on the ST-40 and its mutant at asparagine 255 indicated that the Km value for CS was significantly increased in the mutant without any change in the Km values for 3'-phosphoadenosine 5'-phosphosulfate and DHEA. Inhibition studies demonstrated that DHEA-ST activity was competitively inhibited by AD, but not by CS in the ST-40 enzyme, whereas triethylamine, a noncompetitive inhibitor of hydroxysteroid ST, inhibited DHEA-ST activity in the ST-40 enzyme but did not inhibit CS-ST activity in either ST-40 or ST-20 enzymes. These data provide evidence that DHEA and CS bind to different sites, which probably function in a different manner in the ST-40 enzyme.

Alternative signaling mechanism of leukemia inhibitory factor responsiveness in a differentiating embryonal carcinoma cell.

Leukemia inhibitory factor (LIF) is a cytokine that plays an important role during mouse embryogenesis. We showed that adenovirus E1A represses the interleukin-6 signal transduction pathway that uses the same JAK tyrosine kinase and STAT (signal transducer and activator of transcription) transcription factor as LIF. Here, we report that the LIF-JAK-STAT signal transduction pathway is blocked in cellular E1A-expressing undifferentiated F9 cells, and that the block is overcome by retinoic acid-induced differentiation. LIF failed to stimulate the expression of the acute phase response element (APRE)-driven luciferase gene in undifferentiated F9 cells, whereas the luciferase activity was remarkably increased by LIF treatment in differentiated F9 (dF9) cells. We analyzed the mechanism of the APRE regulation and found that the LIF-induced APRE-binding activity was regulated in a differentiation-dependent manner. The protein levels and the tyrosine phosphorylation of JAK1, JAK2, and STAT3 in F9 cells were not different from those in dF9 cells. The exogenous expression of activated c-Ha-ras partially recovered the LIF responsiveness of the APRE-luciferase gene in F9 cells, but the dominant negative ras N-17 did not repress the LIF-induced activation of APRE-luciferase in dF9 cells. These results suggested that an unknown coactivation process that is partially compensated by Ras is required for STAT3-APRE binding in F9 cells.

Epidermal growth factor promotes adipogenesis of 3T3-L1 cell in vitro.

We have reported the importance of epidermal growth factor (EGF) for the induction of obesity in mice. In this study, we studied the effects of EGF on the induction of lipogenic enzymes and on the accumulation of triglyceride in a differentiated mouse adipocyte cell in vitro. Mouse 3T3-L1 preadipocytic cells differentiated into mature adipocytes after the differentiation procedure by insulin, dexamethasone, and methyl-isobutyl-xanthine. 125I-EGF binding studies in the differentiated 3T3-L1 cells showed specific 125I-EGF bindings, and they expressed gene transcripts for EGF receptors by reverse transcription and polymerase chain reaction at all differentiative stages examined. Although EGF showed inhibitory effects on the triglyceride accumulation when administered to the preadipocytic 3T3-L1 cells, EGF enhanced the adipogenesis in the differentiated cells in dose- and time-dependent manners. Administration of EGF at 0.1-1 nM from 4 days after the differentiation procedure for 10 days, significantly enhanced the acyl-Co A synthetase and lipoprotein lipase messenger RNA levels, both of which are rate-limiting enzymes to synthesize triglyceride in adipocytes. Moreover, 0.1-1 nM EGF increased the amounts of triglyceride accumulated in the cells, in proportion to the acyl-Co A synthetase and lipoprotein lipase messenger RNA levels. EGF rather failed the adipogenesis at 10 nM. Time course studies revealed that 1 nM EGF significantly increased the intracellular triglyceride levels from 4 through 16 days administration. These results suggest that EGF shows biphasic effects on adipocytes: although EGF inhibits preadipocytes differentiation into mature adipocytes, it promotes adipogenesis in the differentiated adipocytes.

Estrogen induces epidermal growth factor (EGF) receptor and its ligands in human fallopian tube: involvement of EGF but not transforming growth factor-alpha in estrogen-induced tubal cell growth in vitro.

We studied the estrogen-dependent expression of epidermal growth factor (EGF), transforming growth factor (TGF) alpha, and EGF receptor gene transcripts in human fallopian tubes in vivo and in vitro. Competitive polymerase chain reaction (PCR) was performed on the fallopian tube RNA samples from the postmenopausal women with or without estrogen replacement. Amounts of EGF, TGF alpha, and EGF receptors gene transcripts in the estrogen-treated group (n = 3) were significantly (P < 0.01) more than those in the untreated group (n = 3). Competitive PCR also showed that EGF, TGF alpha, and EGF receptor gene transcripts level in tubal cells were increased by estrogen in vitro: messenger RNA levels of these factors were significantly (P < 0.01, n = 3) increased in cells incubated with 10(-8) M estrogen compared with those in cells without estrogen treatment. We studied whether EGF and/or TGF alpha is involved in the estrogen-induced tubal cell growth in vitro. Estrogen enhanced the [3H]-thymidine incorporation into the cell in dose- and time-dependent manners in culture: estrogen treatment for more than 12 h significantly (P < 0.05) enhanced the [3H]-thymidine incorporation into the cell at 10(-8) M. The estrogen-induced cell growth was observed in association with the increase in EGF, TGF alpha, and EGF receptor messenger RNA levels by estrogen. If the EGF and/or TGF alpha is involved in the cell growth, then the estrogen-induced cell growth should be suppressed by blocking the action of EGF and/or TGF alpha. Therefore, we examined the effects of neutralizing monoclonal antibodies against EGF, TGF alpha, and EGF receptors. Anti-EGF antibody significantly reduced the estrogen-induced increase in [3H]-thymidine incorporation, whereas anti-TGF alpha antibody failed to show the effect. Anti-EGF receptor antibody showed a significant suppressive effect on the estrogen-induced increase in [3H]-thymidine incorporation. Moreover, the growth inhibitory effect by 1 microgram/ml anti-EGF was restored by 10(-8) M EGF but not by TGF alpha even at 10(-6) M. All these data suggest that estrogen induces EGF and TGF alpha/EGF receptors in the human fallopian tube and that EGF but not TGF alpha may be involved in the estrogen-induced human tubal cell growth in vitro.

Stimulation of steroidogenic acute regulatory protein (StAR) gene expression by D-aspartate in rat Leydig cells.

D-aspartate and human chorionic gonadotropin act synergistically to increase testosterone production in purified rat Leydig cells, and D-aspartate stimulates testosterone synthesis even in the absence of human chorionic gonadotropin stimulation. In addition, D-aspartate enhances steady-state cellular mRNA and protein levels of steroidogenic acute regulatory protein, which is a key regulatory factor in gonadal and adrenal steroidogenesis. D-aspartate therefore appears to increase testosterone production in rat Leydig cells by stimulating steroidogenic acute regulatory protein gene expression. To our knowledge, this is the first report demonstrating a direct effect of D-aspartate on gene expression in mammalian cells.

D-Aspartate stimulation of testosterone synthesis in rat Leydig cells.

D-Aspartate increases human chorionic gonadotropin-induced testosterone production in purified rat Leydig cells. L-Aspartate, D-,L-glutamate or D-,L-asparagine could not substitute for D-aspartate and this effect was independent of glutamate receptor activation. Testosterone production was enhanced only in cells cultured with D-aspartate for more than 3 h. The increased production of testosterone was well correlated with the amounts of D-aspartate incorporated into the Leydig cells, and L-cysteine sulfinic acid, an inhibitor of D-aspartate uptake, suppressed both testosterone production and intracellular D-aspartate levels. D-Aspartate therefore is presumably taken up into cells to increase steroidogenesis. Intracellular D-aspartate probably acts on cholesterol translocation into the inner mitochondrial membrane, the rate-limiting process in steroidogenesis.

Biosynthesis of D-aspartate in mammalian cells.

In this communication, we demonstrate that D-aspartate (D-Asp) is synthesized in pheochromocytoma cells (PC12). To our knowledge this is the first report of biosynthesis of D-Asp in mammalian cells. Synthesis of D-Asp was demonstrated by its time-dependent accumulation in the cell culture, and by the fact that this accumulation was proportional to the number of inoculated cells. D-Asp in PC12 cells was identified by (i) co-elution with authentic D-Asp on two different HPLC columns, an octadesyl silica column and a Pirkle-type chiral column, (ii) reversed elution order of D-Asp and L-Asp on another Pirkle-type chiral column with an opposite configuration, and (iii) sensitivity to D-Asp oxidase. In the cells the amount of D-Asp was approx. 12-14% of total Asp and no other investigated D-amino acid was detected. The amount of D-Asp did not increase during the culture of mouse 3T3 fibroblasts and human neuroblastoma NB-1 cells. Immunocytochemical staining with anti-D-Asp antiserum demonstrated that D-Asp synthesized is present in the cytoplasm of the cells.

Nucleosides and nucleotides. 112. 2-(1-Hexyn-1-yl)adenosine-5'-uronamides: a new entry of selective A2 adenosine receptor agonists with potent antihypertensive activity.

Chemical modifications of the potent A2 adenosine receptor agonist 2-(1-hexyn-1-yl)adenosine (7, 2-HA) at the 5'-position have been carried out to find more potent and selective A2 agonists. These analogues were evaluated for adenosine A1 and A2 receptor binding affinity in rat brain tissues and antihypertensive effects in spontaneously hypertensive rats (SHR). Among the series of compounds, 2-(1-hexyn-1-yl)adenosine-5'-N-cyclopropyluronamide (16d) had the most potent affinity to the A2 receptor with a Ki of 2.6 nM, which is essentially the same as that of the parent agonist, 2-HA. However, the most selective agonist for the A2 receptor was 2-(1-hexyn-1-yl)adenosine-5'-N-methyluronamide (16b) with a Ki of 11 nM and a 162-fold selectivity. The N-alkyl substituents of 5'-uronamide derivatives did not seem to potentiate the A2 binding affinity but drastically reduced the A1 affinity compared with the parent 2-HA. Therefore, the A1/A2 selectivity was consequently increased. Other 5'-deoxy-5'-substituted derivatives of 2-HA such as the chloro (20), carboxamide (27, 28), sulfonamide (29), urea (30), and thiourea (22) analogues were also prepared. Among these nucleosides, no active compounds with potent or selective affinities to both receptors were found except 20. Although glycosyl conformations and sugar-puckering of these nucleosides were studied by 1H NMR spectroscopy, there were no positive correlations between active and inactive agonists. 2-(1-Hexyn-1-yl)adenosine-5'-uronamide (16a) and 16d had a potent hypotensive effect at ED30 values of 0.18 and 0.17 micrograms/kg, respectively, upon iv administration to anesthetized SHR.

Nucleosides and nucleotides. 103. 2-Alkynyladenosines: a novel class of selective adenosine A2 receptor agonists with potent antihypertensive effects.

The synthesis and receptor-binding activities at A1 and A2 adenosine receptors for a series of 2-alkynyladenosines are described. The palladium-catalyzed cross-coupling reaction of 2-iodoadenosine (4a) with various terminal alkynes in the presence of bis(triphenylphosphine)palladium dichloride and cuprous iodide in N,N-dimethylformamide containing triethylamine gives 2-alkynyladenosines (5a-r). An economical synthetic method for the preparation of 9-(2,3,5-tri-O-acetyl-1-beta-D-ribofuranosyl)-6-chloro-2-iodopurine++ + (2), which is a precursor of 4a, is also included. Several transformation reactions of 2-(1-octyn-1-yl)adenosine (5e) and 2-(1-ethyn-1-yl)adenosine (9) and a similar cross-coupling reaction of 6-chloropurine derivative 11 and 8-bromoadenosine (13) with 1-octyne are also reported. Many of these 2-alkynyladenosines tested for A1 and A2 adenosine receptor binding activities in rat brain are selective for the A2 adenosine receptor. Among them, 2-(1-hexyn-1-yl)adenosine (5c) has the highest affinity for both A1 and A2 receptors with Ki values of 126.5 and 2.8 nM, respectively. The structure-activity relationship of this series of compounds including 6- or 8-alkynylpurine nucleosides and 2-alkyl- and 2-alkenyladenosines is discussed in terms of potency at both receptor subtypes. Additionally, we describe how hypotensive activity and heart rate decrease brought on by 5 and some other compounds with spontaneously hypertensive rats are proportional to the order of the potency to both A1 and A2 binding affinities. Thus, 2-alkynyladenosines are interesting and promising as antihypertensive agents that should be considered for further detailed preclinical evaluation.