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The progression of neuroscience relies on the discovery of structure in the brain. From the discovery of neurons to the structure of the potassium channel, and, in recent years, the repeated observation of remarkable geometric structure in the distributed activity of neural populations. What this population-level structure does is not written on it for anyone to read, generally speaking; many statistical and theoretical tools have had to be developed for interpretation. In these chapters, I benefit from and contribute to the growing set of tools for parsing geometries. First, my collaborators and I studied the representation of syntax in (at the time) state-of-the-art language models. Second, we sought to understand why certain geometries emerge in artificial networks. Third, we model the geometry of working memory representations to try and find why 'swap errors' occur. Finally, we offer a new framework and method for discovering discrete structure in continuous representations.

Yes / In a previous study we identified an extensive gating network within the inwardly rectifying Kir1.1 (ROMK) channel by combining systematic scanning mutagenesis and functional analysis with structural models of the channel in the closed, pre-open and open states. This extensive network appeared to stabilize the open and pre-open states, but the network fragmented upon channel closure. In this study we have analyzed the gating kinetics of different mutations within key parts of this gating network. These results suggest that the structure of the transition state (TS), which connects the pre-open and closed states of the channel, more closely resembles the structure of the pre-open state. Furthermore, the G-loop, which occurs at the center of this extensive gating network, appears to become unstructured in the TS because mutations within this region have a 'catalytic' effect upon the channel gating kinetics. / Deutsche Forschungsgemeinschaft, the Wellcome Trust (083547/ Z/07/Z and 092970/Z/10/Z) and the British Heart Foundation (PG/09/016/ 26992).

Kattaya-Annappa-Seema. / Thesis submitted in: December 2006. / "2+" and "+" in the title are superscripts. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2007. / Includes bibliographical references (leaves 162-188). / Abstracts in English and Chinese. / Abstract --- p.i / Acknowledgements --- p.viii / Abstracts published based on work in this thesis --- p.ix / Table of contents --- p.x / Chapter Chapter 1: --- Introduction / Chapter 1.1 --- Pulmonary hypertension / Chapter 1.1.1 --- Pulmonary circulation and its functions --- p.1 / Chapter 1.1.2 --- Pulmonary vascular diseases and symptoms --- p.3 / Chapter 1.2 --- Muscarinic Receptor functions --- p.5 / Chapter 1.3 --- Acetylcholine (ACh) and its function --- p.7 / Chapter 1.4 --- ACh receptors in pulmonary vascular bed --- p.11 / Chapter 1.5 --- Potassium channel classification and functions --- p.12 / Chapter 1.5.1 --- "Importance of High-conductance, Ca2+ activated potassium channel (BKca) in vascular smooth muscle functions" --- p.15 / Chapter 1.5.2 --- Modulation of BKca channel by various cations --- p.18 / Chapter 1.6 --- Calcium signaling and homeostasis --- p.20 / Chapter 1.7 --- Role of sodium in hypertension --- p.22 / Chapter 1.8 --- Na+-H+ exchanger (NHE) functions --- p.25 / Chapter 1.9 --- Na+-Ca2+ exchanger (NCX) in vascular smooth muscle cells --- p.29 / Chapter 1.10 --- Spontaneously hypertensive rat (SHR) / Chapter 1.10.1 --- Hypertension in SHR --- p.32 / Chapter 1.10.2 --- BKca in smooth muscle vasculature of SHR --- p.33 / OBJECTIVES OF THE STUDY --- p.34 / Chapter Chapter 2: --- Material and methods / Chapter 2.1 --- Material / Chapter 2.1.1 --- Solutions and Drugs --- p.35 / Chapter 2.1.2 --- Chemicals and Enzymes --- p.39 / Chapter 2.2 --- Methods / Chapter 2.2.1 --- Isolation of single pulmonary arterial smooth muscle cells --- p.40 / Chapter 2.2.2 --- Electrophysiological measurement --- p.42 / Chapter 2.2.3 --- Data analysis --- p.44 / Chapter Chapter 3: --- Receptor-mediated activation of BKca Channel / Chapter 3.1 --- BKCa activation by ACh/ Carbachol (CCh) --- p.45 / Chapter 3.2 --- Role of extracellular sodium ([Na+]o）on BKca activation --- p.49 / Chapter 3.3 --- Receptor-mediated activation of BKca in a [Na+]o-containing solution --- p.51 / Chapter 3.4 --- Receptor-mediated activation of BKca in a [Na+]o-free solution --- p.55 / Chapter Chapter 4: --- Non-receptor mediated activation of BKCa Channel / Chapter 4.1 --- Effect of different concentrations of sodium nitroprusside (SNP) on BKCa activation --- p.60 / Chapter 4.2 --- Effect of SNP on BKca activation in a [Na+]o-containing and [Na+]o-free solutions --- p.62 / Chapter Chapter 5: --- Role of NHE in modulating activation of BKCa Channel / Chapter 5.1 --- Effect of Monensin on BKca activation / Chapter 5.1.1 --- Effect of monensin on CCh-mediated activation of BKca in a [Na+]o-containing solution --- p.70 / Chapter 5.1.2 --- Effect of monensin on CCh-mediated activation of BKca in a [Na+]o-free solution --- p.74 / Chapter 5.1.3 --- Effect of monensin on SNP- mediated activation of BKca in [Na+]o-containing and [Na+]o-free solutions --- p.78 / Chapter 5.2 --- Effect of 5-(N-ethyl-N-isopropyI) amiloride (EIPA) on BKCa activation / Chapter 5.2.1 --- Effect of EIPA on CCh-mediated activation of BKca in a [Na+]o-containing solution --- p.85 / Chapter 5.2.2 --- Effect of EIPA on CCh-mediated activation of BKca in a [Na+]。-free solution --- p.89 / Chapter 5.2.3 --- Effect of EIPA on SNP-mediated activation of BKCa in [Na+]o-containing and [Na+]o-free solutions --- p.93 / Chapter Chapter 6: --- Role of NCX in modulating activation of BKCa Channel / Chapter 6.1 --- Effect of KB-R7943 on CCh-mediated activation of BKCa in a [Na+]o-containing solution --- p.100 / Chapter 6.2 --- Effect of KB-R7943 on CCh-mediated activation of BKCa in a [Na+]o-free solution --- p.104 / Chapter 6.3 --- Effect of KB-R7943 on SNP-mediated activation of BKca in [Na+]o-containing and [Na+]o-free solutions --- p.109 / Chapter Chapter 7: --- Effect of intracellular sodium ([Na+]i) on BKCa channel activation / Chapter 7.1 --- Effect of CCh on BKCa channel activation with elevated [Na+]i pipette solution --- p.117 / Chapter 7.2 --- Effect of SNP on BKca channel activation with elevated [Na+]j pipette solution --- p.130 / Chapter Chapter 8: --- Discussion / Chapter 8.1 --- Modulatory effect of ACh and SNP --- p.138 / Chapter 8.2 --- Role of ion exchangers: NHE and NCX in modulating BKca channel function --- p.144 / Chapter 8.3 --- Modulatory effect of elevated [Na+]i on BKca activation --- p.153 / CONCLUSION --- p.161 / References --- p.162

KCNE1 (E1) peptide is the founding member of the KCNE family (1-5), which is a class of type I transmembrane ß-subunits. KCNE1 peptides assemble with and modulate the gating, ion conducting properties and pharmacology of a variety of voltage-gated K+ channel a-subunits, including KCNQ1 (Q1). Mutations that interfere with the function of either E1 and/or Q1 and disrupt the assembly and trafficking of KCNE1- KCNQ1 channel complexes give rise to diseases such as Romano-Ward (RW) and Jervell Lange Nielsen Syndrome (JLNS), two different forms of Long QT Syndrome (LQTS). Using enzymatic deglycosylation assays, immunofluorescence techniques and quantitative cell surface labeling, we showed that KCNE1 peptides are retained in the early stages of the secretory pathway as immaturely N-linked glycosylated proteins. KCNE1 co-assembly with KCNQ1 leads to E1 progression through the secretory pathway and glycan maturation, resulting in cell surface expression. N-linked glycosylation of some membrane proteins is critical for proper folding, co-assembly and subsequent trafficking through the biosynthetic pathway. Previous studies have shown that genetic mutations that disrupt one of the two N-linked glycosylation sites on KCNE family members lead to LQTS (T7I, KCNE1 and T8A, KCNE2) (Schulze-Bahr et al., 1997; Sesti et al., 2000a; Park et al., 2003). Having confirmed that KCNE1 proteins acquire N-linked glycans, we examined the kinetics and efficiency of N-linked glycan addition to KCNE1. We showed that KCNE1 has two distinct N-linked glycosylation sites. The N-terminal sequon is a traditional co-translational site. The internal sequon (which is only ~ 20 residues away from the N-terminal sequon) acquires N-linked glycans primarily after protein synthesis (post-translationally). Surprisingly, mutations that prevent N-glycosylation at the cotranslational site also reduce the glycosylation efficiency of post-translational glycosylation at the internal sequon, resulting in a large population of unglycosylated KCNE1 peptides that are retained in the early stages of the secretory pathway and do not reach the cell surface with their cognate K+ channel. We showed that KCNE1 post-translational N-glycosylation in the endoplasmic reticulum is a cellular mechanism that ensures E1 proteins acquire the maximal number of glycans needed for proper channel assembly and trafficking. Our findings provide a new biogenic mechanism for human disease by showing that the JLNS mutation, T7I, not only inhibits glycosylation of the N-terminal sequon, but also indirectly prevents the glycosylation of the internal sequon, giving rise to a large population of assembly incompetent hypoglycosylated KCNE1 peptides. To further investigate the two N-linked glycosylation sites on KCNE1, we generated structure-function deletion scans of KCNE1 and performed positional glycosylation scanning mutagenesis. We examined the glycosylation pattern of glycosylation mutants in an effort to define the glycosylation window important for proper KCNE1 assembly and trafficking. Our findings suggested a nine amino acid periodicity to serve as a desirable glycosylation site and a better substrate for N-glycosylation. Appendix II shows work on the characterization of the C-terminally HA-tagged KCNE1 protein, which was used throughout the experiments presented in Chapter II, Chapter III and Chapter IV. Analysis of the C-terminally HA-tagged KCNE1 protein revealed that in heterologous expression systems KCNE1 had an internal translational start site, a methionine at position 27. A proteolytic cleavage site was also identified at the arginine cluster spanning residues 32 through 38 bearing the two known Long QT mutations (R32H and R36H) (Splawski et al., 2000; Napolitano et al., 2005). My work in Professor Craig C. Mello’s lab during the first four years of my graduate study is presented in Appendix I. The highly conserved Wnt/Wingless glycoproteins regulate many aspects of animal development. Wnt signaling specifies endoderm fate by controlling the fate of EMS blastomere daughters in 4-cell stage Caenorhabditis elegans embryos. A suppressor genetic screen was performed using two temperature sensitive alleles of mom-2/Wnt to identify additional regulators of the Wnt/Wingless signaling pathway during C. elegans endoderm specification. Five intragenic suppressors and three extragenic suppressors of mom-2/Wnt embryonic lethality were identified. We cloned ifg-1, eIF4G homologue, as one of the extragenic suppressors suggesting an intriguing connection between the Wnt signaling pathway and the translational machinery.

O diabete melito é um grupo de doenças metabólicas caracterizadas por hiperglicemia, resultado de deficiências na secreção de insulina, em sua acção ou ambos. A pioglitazona é um hipoglicemiante oral, da classe da tiazolidinedionas, que atuam pela ligação aos receptores nucleares PPARY melhorando o estado de resistência a insulina. Acredita-se que a pioglitazona também restauram a capacidade da célula beta pancreática de secretar insulina, cuja atividade é regulada pelos canais de potássio dependente de ATP (KATP) e suas subunidades SUR1 e KIR6.2. Este estudo teve como objetivo iniciar um estudo farmacogenômico da pioglitazona em indivíduos diabéticos tipo 2 e na expressão dos genes PPARY PPARY2, SUR1 e KIR6.2 no sangue periférico e no tecido adiposo e associá-los com os polimorfismo Pro12Ala e C161T do gene PPARY. Foram selecionados 36 pacientes diabéticos do tipo 2 e 16 pacientes normoglicêmicos, no Instituto Dante Pazzanese de Cardiologia. Os indivíduos diabéticos foram tratados com pioglitazona (15, 30 e 45 mg/ dia/ via oral) por 16 semanas. Foram colhidas amostras de tecido adiposo por biopsia e de sangue, antes e após o tratamento para determinação de exames laboratoriais, extração de DNA genômico e de RNA total. Os polimorfismos foram detectados pela técnica de PCR-RFLP e a expressão de mRNA foi quantificada e avaliada por RT-PCR em tempo real. Após tratamento com pioglitazona, observou-se no sangue periférico aumento de expressão de mRNA do PPARY, PPARY2 e KIR6.2, e diminuição da expressão de SUR1. Dados de analises no sangue periférico, demostraram que variação da expressão de mRNA do PPARY foi inversamente correlacionada com as variações de insulina, Homa-IR, Homa-Beta e positivamente com Colesterol Total. Em relação ao gene PPARY2, foi inversamente correlacionada com colesterol total. Variação da expressão de SUR1 foi inversamente correlacionada com Hb1Ac, Homa-IR, Homa-Beta e positivamente com insulina. Não foram detectadas diferenças entre a expressão de mRNA de KIR6.2 e parâmetros bioquímicos em resposta às variações pioglItazona. Não houve diferença da resposta terapêutica e presença dos polimorfismos Pro12Ala e C161T. Após o tratamento, no tecido adiposo, a expressão de mRNA de PPARY aumentou. Não foram observadas diferenças na expressão dos genes PPARY2, SUR1 e KIR6.2. Em leucócitos totais de sangue periférico, pioglitazona demonstrou regular a expressão dos genes PPARY e PPARY2, e também atuando sobre as subunidades SUR1 e KIR6.2, possivelmente restaurando a função secretora das células beta. No tecido adiposo, o tratamento confirma a atuação da pioglitazona sobre PPARY, melhorando o estado de resistência a insulina. / The diabetes mellitus is a group of metabolic diseases characterized by hyperglycemia, the result from insulin secretion or action deficiency, or both. Pioglitazone is an oral hypoglycemic drug, included in the class of thiazolidinediones, which work by binding to nuclear receptors PPARY nuclear what improve the state of resistance to insulin. Pioglitazone may also restore the ability of the pancreatic beta cell to secrete insulin, process which is regulated thought ATP dependent potassium channels (KATP) and its subunits SUR1 and KIR6.2. The aim of our study was to begin a pharmacogenomic study of pioglitazone in type 2 diabetes patients by the expression of PPARY, PPARY2, SUR1 and KIR6.2 genes in peripheral blood leukocytes and fatty tissue and its association with the PPARY Pro12Ala and C161T polimorphisms. 36 type 2 diabetes and 16 normoglycemic patients were selected at the Dante Pazzanese Institute of Cardiology. The diabetic ones were treated with pioglitazone (15, 30 and 45 mg/ daily /orally) for 16 weeks. Samples of adipose - obtained through biopsy - and blood were collected before and after treatment to aim to laboratory experiments, DNA extraction and total RNA extraction. The polymorphisms were detected by the PCR-RFLP technique while the mRNA expression was quantified and evaluated by Real Time RT-PCR. After pioglitazone treatment, the expression of the PPARY, PPARY2 and KIR6.2 genes increased, while SUR1 decreased, all of them quantified in peripheral blood. Peripheral blood data has demonstrated that the variation of expression of PPARYmRNA is inversely correlated with insulin concentrations, Homa-IR, Homa-Beta and positively correlated with total cholesterol concentrations. Differently, PPARY2 gene expression was inversely correlated with total cholesterol blood concentrations. Variations in SUR1 mRNA expression were inversely correlated with Hb1Ac, Homa-IR, Homa-Beta and positively correlated with insulin. No differences were found between the KIR6.2 expression and biochemical parameters due to pioglItazone treatment. There was no difference in response to the treatment response and no Pro12Ala and C161T polymorphisms were noticed. After the treatment, the expression of PPARY gene increased in fatty tissue. No data showed differences in the expression of genes PPARY2, SUR1 and KIR6.2. Pioglitazone regulates the expression of genes PPARY and PPARY2, and also acts on the SUR1 and KIR6.2 subunits in total peripheral blood leukocyte likely restoring the function of secreting beta pancreatic cells. In adipose, the treatment reassures the functions of pioglitazone on PPARY, improving the state of insulin resistance.

O hiperinsulinismo congênito (CHI) é um distúrbio do pâncreas endócrino, mais freqüentemente causado por alterações dos canais de membrana KATP das células , resultando em secreção inapropriada de insulina e hipoglicemia severa e persistente nos recém-nascidos, que leva ao óbito ou a seqüelas neurológicas graves, se não diagnosticado a tempo. O diagnóstico depende da análise dos dados clínicos, laboratoriais, morfológicos e genético-moleculares (50% apresentam mutações dos canais KATP). As duas formas histopatológicas descritas requerem cirurgias radicalmente opostas: pancreatectomia quase-total (95-98%) na forma difusa que acomete todo o pâncreas, ou apenas exerese do foco adenomatoso de células , medindo em média 4,5 mm, na forma focal, e portanto a sua distinção é essencial durante o exame intra-operatório de congelação ou através de [18F]-L-Dopa PET-CT. Dez pacientes com CHI difuso e um com CHI focal, submetidos a pancreatectomia, foram analisados em relação a parâmetros clínicos, histopatológicos, de proliferação das células (IHQ de dupla marcação Ki-67 / insulina) e quanto à presença de mutações nos genes das únicas duas proteínas (SUR 1 e Kir 6.2) que formam os canais KATP, e comparados a 19 pâncreas controles normais da mesma faixa etária. Pacientes e controles foram estratificados em 3 meses e > 3 meses de idade. Nucleomegalia, ausente nos controles, foi observada apenas na forma difusa. Os critérios histológicos de maturação normalmente mais freqüentes nos controles 3 meses, foram freqüentemente observados nos recém-nascidos com CHI difuso > 3 meses, sugerindo um retardo na maturação do pâncreas endócrino destes pacientes. O índice de proliferação das células (Ki-67- LI), muito elevado nos focos adenomatosos da forma focal, foi útil na distinção destes focos dos agregados frouxos de ilhotas, histologicamente muito semelhantes, observados em dois casos difusos e um controle, que apresentam níveis de Ki-67-LI cerca de 10 vezes menor. Na forma difusa o Ki-67-LI também foi estatisticamente mais alto do que nos controles. Este é o primeiro estudo de pacientes com CHI no Brasil, e embora existam diferenças epidemiológicas entre os países relacionadas à determinação genética do CHI, não foram constatadas mutações ou novos polimorfismos nos exons 33-37 do gene ABCC8 (SUR 1) de 10/10 pacientes ou no único exon do gene KCNJ11 (Kir 6.2) de 4/10 pacientes / Congenital hyperinsulinism (CHI) is a rare pancreatic endocrine cell disease which most severe cases are found to be, at least in half of patients, associated with genetic defects in the -cell KATP channels. The aim of this study was to evaluate eleven Brazilian patients diagnosed, by standard criteria, as CHI non responsive to clinical therapy, and submitted to pancreatectomy, regarding: histology, -cell proliferation (IHC Ki-67 / insulin) and -cell KATP channels genes mutations in blood samples. For comparison of histology and -cell proliferation, 19 pancreatic control samples were included. According histology, ten patients were classified as diffuse and one as focal form. Nucleomegaly and -cells with abundant cytoplasm were absent in controls, and observed only in the group of diffuse CHI patients. Ki- 67-LI was useful to differentiate the adenomatous areas of the focal form CHI neonate from loose clusters of islets found in two diffuse form and one control samples. Proliferation was much higher in the focal CHI adenomatous areas, but diffuse CHI patients also have statistically higher Ki-67-LI than controls. This is the first genetic study of CHI patients in Brazil, and no mutations or new polymorphisms were found in the ABCC8 gene (SUR 1) (exons 33-37) or in the only exon of KCNJ11 gene (Kir 6.2) in 4/4 patients evaluated. On the other hand, enhanced -cell proliferation seems to be a constant feature in these patients both in diffuse and focal forms

The voltage gated potassium channel Kv1.2 plays a key role in the central nervous system and mutations in Kv1.2 cause neurological disorders such as epilepsies and ataxias. In the cerebellum, regulation of Kv1.2 is coupled to learning and memory. We have previously shown that blocking Kv1.2 by infusing its specific inhibitor tityustoxin-kα (TsTX) into the lobulus simplex of the cerebellum facilitates eyeblink conditioning (EBC) and that EBC itself modulates Kv1.2 surface expression in cerebellar interneurons. The metabotropic glutamate receptor mGluR1 is required for EBC although the molecular mechanisms are not fully understood. Here we show that infusion of the mGluR1 agonist (S)-3,5-dihydroxyphenylglycine (DHPG) into the lobulus simplex of the cerebellum mimics the facilitating effect of TsTX on EBC. We therefore hypothesize that mGluR1 could act, in part, through suppression of Kv1.2. Earlier studies have shown that Kv1.2 suppression involves channel tyrosine phosphorylation and endocytocytic removal from the cell surface. In this study we report that an excitatory chemical stimulus (50mM K+-100µM glutamate) applied to cerebellar slices enhanced Kv1.2 tyrosine phosphorylation and that this increase was lessened in the presence of the mGluR1 inhibitor YM298198. More direct evidence for mGluR1 modulation of Kv1.2 comes from our finding that selective activation of mGluR1 with DHPG reduced the amount of surface Kv1.2 detected by cell surface biotinylation in cerebellar slices. To determine the molecular pathways involved we used an unbiased mass spectrometry-based proteomics approach to identify Kv1.2-protein interactions that are modulated by mGluR1. Among the interactions enhanced by DHPG were those with PKC-γ, CaMKII, and Gq/G11, each of which had been shown in other studies to co-immunoprecipitate with mGluR1 and contribute to its signaling. Of particular note was the interaction between Kv1.2 and PKC-γ since in HEK cells and hippocampal neurons Kv1.2 endocytosis is elicited by PKC activation. We found that activation of PKCs with PMA reduced surface Kv1.2, while the PKC inhibitor Go6983 attenuated the reduction in surface Kv1.2 levels elicited by DHPG and PMA, suggesting that the mechanism by which mGluR1 modulates cerebellar Kv1.2 likely involves PKC. mGluR1 has been shown to signal independently of the agonist through a constitutively active, protein kinase A-dependent pathway in the cerebellum. Using HEK293 cells we show that co-expression of mGluR1 increases the surface expression levels of Kv1.2. This effect occurs in absence of mGluR1 agonists and in the presence of a noncompetitive mGluR1 inhibitor YM298198. Co-expression of known downstream effectors of the agonist driven mGluR1 pathway such as PKC-γ, CaMKIIα, Grid2 had no effect on Kv1.2 surface expression or on the ability of mGluR1 agonist to modulate that expression. In contrast, the inverse agonist BAY 36-7620 significantly reduced the mGluR1 effect on Kv1.2 surface expression, as did pharmacological inhibition of PKA with KT5720. Therefore, mGluR1 is involved in regulation of surface Kv1.2 via dual mechanisms, the agonist dependent mechanism reduces surface Kv1.2 via PKC, while agonist independent constitutive mechanism increases surface Kv1.2 via PKA.

Eine optimale Regulation der Koronardurchblutung ist für die Aufrechterhaltung der kardialen Pumpfunktion und damit der systemischen Perfusion von größter Bedeutung. Da Einschränkungen der Durchblutungszunahme des Herzmuskels Einschränkungen des maximalen myokardialen Sauerstoffverbrauchs und damit der Herzleistung zur Folge haben, ist es notwendig, die Koronardurchblutung kurzfristig an die jeweilige Stoffwechsellage des Herzens anzupassen (metabolische Koronarflussregulation). Die lokal-metabolischen Mechanismen gehören zu den wirksamsten Komponenten der Regulation der myokardialen Durchblutung und funktionieren auch am isolierten (denervierten) Herz. Dabei ist die hyperkapnie- und azidoseinduzierte Koronardilatation ein wesentlicher Bestandteil der metabolischen Koronarflussregulation. Die vorliegende Arbeit befasst sich mit der Hypothese der Abhängikeit der hyperkapnieinduzierten Koronardilatation von einer intakten NO-Produktion. Das Koronarsystem des isolierten WT-Mausherzens reagiert auf akute Hyperkapnie (91 % O2, 9 % CO2) mit einer deutlichen Koronarflusssteigerung von ca. 35 % über dem Basalfluss.Es konnte gezeigt werden das Stickstoffmonoxid (NO) und ATP-abhängige Kaliumkanäle (K+ATP-Kanäle) für die Koronarflussregulation der Maus eine ausschlagebende Rolle spielen und neben der Aufrechterhaltung des Basalflusses auch an der Vermittlung der hyperkapnieinduzierten Koronardilatation maßgeblich beteiligt sind.Interessanterweise ist bei einem Fehlen der endothelialen NO-Synthase durch genetischen Knockout die hyperkapnieinduzierte Flussantwort in Kinetik und Ausmaß vollständig erhalten. Die Vermittlung kann dabei durch andere Mechanismen kompensiert werden, wie zum Beispiel einer verstärkten Aktivität der K+ATP-Kanäle. Prostaglandine und neuronale NO-Synthase scheinen sowohl beim Wildtypherzen als auch bei Herzen mit fehlender NO-Synthase für die hyperkapnieinduzierte Koronardilatation von untergeordneter Bedeutung. Nach chronischer pharmakologischer Blockade der NO-Synthase durch zweiwöchige L-NAME Tränkung bleibt die hyperkapnieinduzierte Koronardilatation erhalten durch NOS-unabhängige Mechanismen. Die hyperkapnieinduzierte Flussantwort ist bei Herzen von weiblichen eNOSKO Tieren vorhanden, erscheint jedoch gegenüber den männlichen Mäusen geringer ausgeprägt. Daher wird vermutet, dass die Mediatorsysteme der endothelabhängigen Koronarflussregulation geschlechtsspezifisch bzw. geschlechtsabhängig sind.