Xiao-Fang Li

K+-dependent Na+/Ca2+ exchangers in the brain.

Abstract: Sodium‐calcium exchange was first characterized in heart myocytes and squid axon more than 3 decades ago. Since then, it has been appreciated that functioning of the Na/Ca exchanger molecule plays a critical role in calcium homeostasis in neurons. Genome analysis indicates that Na/Ca exchangers are a superfamily encoded by 7 different genes divided into 2 groups: the Na/Ca exchangers (NCX; SLC8) and the Na/Ca+K exchangers (NCKX; SLC24). Two different NCX genes, NCX1 and NCX2, are highly expressed in brain. We recently described the widespread expression of 2 NCKX‐type exchangers in brain, NCKX2 and NCKX3, and uncovered evidence for expression of another, NCKX4. The unique role that each different exchanger plays in neuronal calcium homeostasis, however, awaits further investigation. To begin exploring this central question, we examined both the expression pattern and the functional properties of the K‐dependent Na/Ca exchanger isoforms expressed in brain and compared and contrasted these with NCX‐type exchangers. Distinct patterns of transcript abundance, regional distribution, and developmental expression were noted for each isoform. Functional properties, including stoichiometry and the kinetic characteristics of ion binding, were determined for NCKX2 and are discussed in the context of cellular Ca2+ signaling.

Functional characterization and m-RNA expression of 5-HT receptors mediating contraction in human umbilical artery

5‐HT1‐like and 5‐HT2 receptors have both been described to mediate contractions to 5‐HT in the human umbilical artery (HUA). However, the nature of the 5‐HT receptor subtypes is unknown. In isometric force studies with ring preparations of HUA α‐methyl‐5‐hydroxytryptamine (α‐Me‐5‐HT) and 5‐hydroxytryptamine (5‐HT) contracted HUA with pED50 values of 8.04 and 7.74, respectively. In the presence of a subthreshold concentration of another vasoconstrictor sumatriptan and 5‐nonyloxytryptamine elicited concentration‐dependent contractions with pEC50 values of 7.21 and 7.67, respectively. In the presence of the selective 5‐HT1B/D receptor antagonist GR127935, contractile responses elicited by sumatriptan and 5‐nonyloxytryptamine were competitively antagonized (pKB 9.01 and 9.02, respectively). In the experiments with 5‐HT, GR127935 appeared to be non‐competitive with shallow Schild plot slopes. The data were fitted with two linear regression lines and the calculated pKB of the high affinity component (8.90) was comparable to that expected for GR127935 at the 5‐HT1B/1D receptor. Several 5‐HT2 selective receptor antagonists (spiperone, cyproheptadine, pirenperone) competitively inhibited responses to 5‐HT. The selective 5‐HT2A antagonist ketanserin against sumatriptan and 5‐nonyloxytryptamine behaved as a weak antagonist while against 5‐HT demonstrated a competitive antagonism (pKB 8.56). Using specific primers for human 5‐HT1B, 5‐HT1D and 5‐HT2A receptor genes, the reverse transcriptase‐polymerase chain reaction revealed mRNA expression of 5‐HT1B and 5‐HT2A receptors in the HUA. The results suggest that the HUA has a functional population of 5‐HT1B and 5‐HT2A receptor subtypes which are involved in the contractile response to 5‐HT. Contractions mediated by 5‐HT1B receptors can be ‘uncovered’ by exposure to other vasoactive agents.

The effects of α-adrenoceptor agonists on intracellular Ca2+ levels in freshly dispersed single smooth muscle cells from rat tail artery

1 The presence of functional α‐adrenoceptors in freshly dispersed single smooth muscle cells from rat tail arteries was investigated by use of selective α‐adrenoceptor agonists and antagonists. 2 Cirazoline, a selective α1‐adrenoceptor agonist, caused a prazosin‐sensitive, rapid but transient increase in intracellular Ca2+, which was partially inhibited by the voltage‐dependent Ca2+ channel blocker, nifedipine. 3 TL99, an α2‐adrenoceptor agonist, in the presence of prazosin, initiated a slow and sustained increase in intracellular Ca2+ which was partially inhibited by yohimbine and almost completely blocked by nifedipine. 4 In rat tail artery, the effects (dose‐response and time‐response curves) of cirazoline and TL99 on intracellular Ca2+ levels in freshly dispersed single smooth muscle cells were comparable with those obtained with organ bath studies of ring preparations of artery. 5 In freshly dispersed single smooth muscle cells, the time‐course response curves induced by the selective α1‐adrenoceptor agonist, phenylephrine and the selective α2‐adrenoceptor agonist, UK14304, were similar to those observed with cirazoline and TL99, respectively. 6 These results indicate that: (a) functional α1‐ and α2‐adrenoceptors are present in freshly dispersed single smooth muscle cells from rat tail artery and (b) α1‐ and α2‐adrenoceptors are coupled to different cellular processes that lead to an increase in intracellular Ca2+.

Tension Development, Myosin Light Chain Phosphorylation and Changes in Intracellular Free [Ca2+]i Induced by α1- and α2- Adrenoceptor Agonists

Vascular smooth muscle tone is regulated by a complex set of processes which vary with location of the vascular bed and the species being studied. Thus, in any synthesis of these regulatory processes, one must consider not only the effects of neurotransmitters but also vasoactive factors including hormones, locally produced substances (e.g., endothelium-derived factors) and changes in the local ionic milieu. Much emphasis has been placed on describing the effects of a specific vasoactive substance, or class of substances, on a particular vascular tissue. Much less emphasis, however, has been placed on understanding the cellular basis for the interactions (synergistic or otherwise) which occur between different vasoactive substances. To address this deficit, we are investigating the roles of α1- and α2- adrenoceptors in the regulation of vascular tone. The overall objective of this study was to determine not just the effects of individual agonists but to analyse the synergistic interactions that occur when vascular smooth muscle cells are exposed to sub-threshold concentrations of angiotensin II (ATII) prior to α-adrenergic agonists.

Signal Transduction in Vascular Smooth Muscle: Synergistic Interactions Between Receptors and Transduction Pathways

Angiotensin II (AII) is an extremely potent vasoconstrictor that is derived from angiotensin via the enzymic action of renin to first yield angiotensin I and then, via cleavage by angiotensin converting enzyme (ACE), AII. ACE inhibitors (ACEIs) are well established as important therapeutic agents for treating both hypertension and congestive heart failure; therapeutic efficacy is not clearly related to plasma renin or AII levels, suggesting “non-selective” actions of ACEIs and/or that plasma renin/AII levels do not reflect the true physiological/pathophysiological function of AII in the vasculature.