Barbara J. Elmquist

Expression of Shal potassium channel subunits in the adult and developing cochlear nucleus of the mouse

The pattern of expression of potassium (K(+)) channel subunits is thought to contribute to the establishment of the unique discharge characteristics exhibited by cochlear nucleus (CN) neurons. This study describes the developmental distribution of mRNA for the three Shal channel subunits Kv4.1, Kv4.2 and Kv4.3 within the mouse CN, as assessed with in situ hybridization and RT-PCR techniques. Kv4.1 was not present in CN at any age. Kv4.2 mRNA was detectable as early as postnatal day 2 (P2) in all CN subdivisions, and continued to be constitutively expressed throughout development. Kv4.2 was abundantly expressed in a variety of CN cell types, including all of the major projection neuron classes (i.e., octopus, bushy, stellate, fusiform, and giant cells). In contrast, Kv4.3 was expressed at lower levels and by fewer cell types. Kv4.3-labeled cells were more prevalent in ventral subdivisions than in the dorsal CN. Kv4.3 expression was significantly delayed developmentally in comparison to Kv4.2, as it was detectable only after P14. Although the techniques employed in this study detect mRNA and not protein, it can be inferred from the differential distribution of Kv4 transcripts that CN neurons selectively regulate the expression of Shal K(+) channels among individual neurons throughout development.

Differential Effects of Complement Activation Products C3a and C5a on Cardiovascular Function in Hypertensive Pregnant Rats

Early-onset pre-eclampsia is characterized by decreased placental perfusion, new-onset hypertension, angiogenic imbalance, and endothelial dysfunction associated with excessive activation of the innate immune complement system. Although our previous studies demonstrated that inhibition of complement activation attenuates placental ischemia–induced hypertension using the rat reduced uterine perfusion pressure (RUPP) model, the important product(s) of complement activation has yet to be identified. We hypothesized that antagonism of receptors for complement activation products C3a and C5a would improve vascular function and attenuate RUPP hypertension. On gestational day (GD) 14, rats underwent sham surgery or vascular clip placement on ovarian arteries and abdominal aorta (RUPP). Rats were treated once daily with the C5a receptor antagonist (C5aRA), PMX51 (acetyl-F-[Orn-P-(D-Cha)-WR]), the C3a receptor antagonist (C3aRA), SB290157 (N2-[(2,2-diphenylethoxy)acetyl]-l-arginine), or vehicle from GD 14–18. Both the C3aRA and C5aRA attenuated placental ischemia–induced hypertension without affecting the decreased fetal weight or decreased concentration of free circulating vascular endothelial growth factor (VEGF) also present in this model. The C5aRA, but not the C3aRA, attenuated placental ischemia–induced increase in heart rate and impaired endothelial-dependent relaxation. The C3aRA abrogated the acute pressor response to C3a peptide injection, but it also unexpectedly attenuated the placental ischemia–induced increase in C3a, suggesting nonreceptor-mediated effects. Overall, these results indicate that both C3a and C5a are important products of complement activation that mediate the hypertension regardless of the reduction in free plasma VEGF. The mechanism by which C3a contributes to placental ischemia–induced hypertension appears to be distinct from that of C5a, and management of pregnancy-induced hypertension is likely to require a broad anti-inflammatory approach.

Neutrophil Depletion Attenuates Placental Ischemia-Induced Hypertension in the Rat.

Preeclampsia is characterized by reduced placental perfusion with placental ischemia and hypertension during pregnancy. Preeclamptic women also exhibit a heightened inflammatory state and greater number of neutrophils in the vasculature compared to normal pregnancy. Since neutrophils are associated with tissue injury and inflammation, we hypothesized that neutrophils are critical to placental ischemia-induced hypertension and fetal demise. Using the reduced uteroplacental perfusion pressure (RUPP) model of placental ischemia-induced hypertension in the rat, we determined the effect of neutrophil depletion on blood pressure and fetal resorptions. Neutrophils were depleted with repeated injections of polyclonal rabbit anti-rat polymorphonuclear leukocyte (PMN) antibody (antiPMN). Rats received either antiPMN or normal rabbit serum (Control) on 13.5, 15.5, 17.5, and 18.5 days post conception (dpc). On 14.5 dpc, rats underwent either Sham surgery or clip placement on ovarian arteries and abdominal aorta to reduce uterine perfusion pressure (RUPP). On 18.5 dpc, carotid arterial catheters were placed and mean arterial pressure (MAP) was measured on 19.5 dpc. Neutrophil-depleted rats had reduced circulating neutrophils from 14.5 to 19.5 dpc compared to Control, as well as decreased neutrophils in lung and placenta on 19.5 dpc. MAP increased in RUPP Control vs Sham Control rats, and neutrophil depletion attenuated this increase in MAP in RUPP rats without any effect on Sham rats. The RUPP-induced increase in fetal resorptions and complement activation product C3a were not affected by neutrophil depletion. Thus, these data are the first to indicate that neutrophils play an important role in RUPP hypertension and that cells of the innate immune system may significantly contribute to pregnancy-induced hypertension.

C-Type Natriuretic Peptide Attenuates Evoked Dopamine Efflux by Influencing Goα

Abstract —Natriuretic peptides suppress adrenergic neurotransmission by a mechanism sensitive to pertussis toxin, suggesting that GTP-binding proteins are involved in the response. The major GTP-binding proteins present in the pheochromocytoma (PC12) cells used in this report are Goα and Giα2. We tested the hypothesis that the more abundant GTP-binding protein, Goα, mediates natriuretic peptide effects in PC12 cells by selectively ablating Goα from the cells with antisense oligodeoxynucleotides. The results indicate that a selective ablation of Goα with this technique eliminated C-type natriuretic peptide (CNP) effects and suppressed dopamine efflux evoked by a depolarizing stimulus. However, the activation of guanylyl cyclase (GC) by CNP was sustained after the Goα ablation. Further, N ω-nitro-l-arginine methyl ester suppressed evoked dopamine efflux equally in the presence and absence of Goα. These results suggest that CNP attenuates evoked catecholamine efflux from PC12 cells by a mechanism requiring Goα but independent of GC activation.

Atrial Natriuretic Factor Binding to Clearance Receptors in Isolated Neuronal Membranes

Publisher Summary Atrial natriuretic factor (ANF) interacts with at least two distinct receptors to produce various biological effects. The two receptors that bind ANF with similar affinities can be differentiated by molecular mass. The larger receptor, termed the R 1 or B receptor, generates cyclic guanosine monophosphate (cGMP) on stimulation with ANF; moreover, cloning studies have identified it as a particulate guanylate cyclase. The smaller receptor is identified as the R 2 or clearance receptor, existing as a monomer or a dimer. The R 2 receptor specifically binds the truncated ANF derivative, ANF–NH 2 (cANF), whereas this analog does not activate the R 1 receptor. Therefore, cANF has been utilized in numerous studies to identify specific ANF binding to the R 1 receptor and to displace binding from the R 2 receptor. Moreover, cANF mimics the effects of ANF on peripheral sympathetic neurons to suppress the evoked catecholamine release and adenylate cyclase activity in a manner similar to ANF. These results suggest that the R 2 receptor is the biologically functional ANF receptor in the peripheral sympathetic nervous system. Methods for the identification of neuronal ANF receptors involve: (1) binding of labeled cANF to membranes, (2) coupling of [ 125 I]ANF to receptors with subsequent identification of R 1 or R 2 receptors by molecular mass, and (3) utilizing cANF to displace selectively ANF binding to the R 2 receptor.