Theme 3: Vascular Biology Flashcards

Question

Summarise how opening and closing of potassium channels affects vasodilation in arterial smooth muscle cells.

Answer 1

* Chloride -\> E_Cl = -20mV * Sodium -\> E_Na = +79mV * Calcium -\> E_Ca = +126mV Each of these will lead to depolarisation when opened, since the resting membrane potential is usually more negative than these equilibrium potentials.

Answer 2

* Arterial smooth muscle cells concentrate chloride using the NKCC and anion exchanger (AE) * This means that the equilibirum potential for chloride is much less negative (around -20mV) than in other cells (usually close to the equilibrium potential of potassium) * This means that opening of chloride channels leads to depolarisation, rather than hyperpolarisation

Answer 3

(Byrne, 1987): * Used carbachol on isolated smooth muscle cells and perfomed patch clamping on them * The carbachol is an agonist of a GPCR that leads to opening of chloride channels * This leads to inward currents * Plotted an I/V graph that shows that the equilibirum potential for chloride here is around 0mV (which is less negative than the usual -20mV) * The downside of using isolated SMCs in solution is that SMCs usually act as a syncytium so this is not representative. Also, it is difficult to control conditions.

Answer 4

* The chloride channels are of two main types: * TMEM16A * Bestrophin * These are both calcium-activated, leading to depolarisation of the membrane -\> This in turn leads to opening of VGCCs and entry of more calcium * This provides an explanation for how GPCRs can lead to opening of chloride channels (by increasing intracellular calcium) * Bestrophin channels are also activated by cGMP, providing another route for modulation

Answer 5

(Heinze, 2014): * Knocked out the calcium-activated chloride channel TMEM16A in vascular SMCs, intermediate cells, and pericytes * This led to elimination of calcium-activated chloride currents and caused lower systemic blood pressure and a decreased hypertensive response following vasoconstrictor treatment. * This suggests that TMEM16A plays a general role in arteriolar and capillary blood flow and is a promising target for the treatment of hypertension.

Answer 6

* Evidence for their involvement is limited, although there is some * (Keatinge, 1968): * Carried out sucrose gap recordings (a technique no longer used) on isolated vascular smooth muscle cells * Used a calcium-free solution where the calcium was replaced with a non-permeant similar ion * Observed sodium-based voltage spikes under these conditions, which were lost when the calcium in the solution was replaced * This suggests that sodium currents could play a role in arterial smooth muscle cells, but are inhibited by physiological concentrations of calcium * (Ho, 2013): * Used veratridine to block Na_v channels in arterial smooth muscle cells, which led to reduced vasoconstriction * It was suggested that Na⁺ influx drives reverse mode NCX, Ca²⁺ entry and Cl^-_Ca activation to cause depolarisation * Another route for Na+ entry is via TRPM4 and possibly other ‘non-selective’ cation channels

Answer 7

* There is no evidence specifically in arterial smooth muscle, but there is evidence from other types of smooth muscle * (Bulbring, 1963): * Studied action potentials in guinea pig taenia coli muscles * Found that these were calcium-based * Their frequency was increased by stretch

Answer 8

* Calcium sparks -\> Spontaneous intracellular release through RyR * Calcium puffs -\> Spontaneous intracellular release through IP₃R Sparks and puffs both interact to generate cell-wide and rapid Ca²⁺ waves. * Calcium sparklets -\> Entry of extracellular calcium through individual VGCCs * TRPV4 sparklets -\> Entry of extracellular calcium through individual TRPV4 channels

Answer 9

1. Direct action -\> Promoting vasoconstriction 2. Indirect -\> Suppressing vasoconstriction

Answer 10

The Bayliss effect (myogenic tone)

Answer 11

* Small resistance arteries and arterioles * This could be explained by a different distribution of ion channels

Answer 12

Calcium sparks: * Calcium sparks activate BK_Ca (calcium-dependent potassium channels) on the membrane, leading to hyperpolarisation * This reduces Ca²⁺ influx via VGCCs, relaxing arterial smooth muscle This is dependent on the specific arrangement of proteins in the smooth muscle cell (since the calcium sparks are very local).

Answer 13

This allows negative feedback modulation of myogenic tone and agonist-driven vasoconstriction.

Answer 14

Mostly the dilation of skeletal muscle microcirculation

Answer 15

1. Mechanisms limiting blood flow: * Myogenic tone * Vasoconstrictors 2. Mechanisms increasing blood flow: * Endothelial cell-dependent mechanisms (myoendothelial feedback) * Metabolic + Conducted vasodilation

Answer 16

* Flow-induced vasodilation is only seen in large arteries * Voltage-dependent processes are seen in smaller vessels

Answer 17

* Myogenic tone is the contraction of small vessels in response to increases in pressure in the lumen * Experimental evidence: * Used an isolated cannulated cremaster muscle arteriole ex vivo * Suddenly increased the pressure from 5mmHg to 80mmHg * This led to an increase in the diameter, followed by a return to baseline diameter * The vessel diameter was then plotted against the luminal pressure at increments from 5 to 80mmHg, both with and without removal of calcium to show active and passive processes

Answer 18

Myogenic tone is driven by depolarisation and calcium entry: * Plotted a graph of % artery diameter against luminal pressure (normalised to 100% at 70mmHg) -\> Plotted a line for the passive diameter (when calcium is removed) against the active diameter (normal) * Also plotted smooth muscle cell membrane potential against luminal pressure * At 75mmHg, there was around 50% tone (i.e. the active diameter was half of the passive diameter) * Nifedipine (L-type calcium channel blocker) blocked the tone but not the depolarisation -\> This suggests that calcium entry is involved in myogenic tone, but L-type calcium channels are not involved in the depolarisation The depolarisation drives increased calcium entry into SMCs and increased phosphorylation of MLC: * Utilised FURA-2 as an indicator of intracellular calcium, which increased proportionally to diameter during the myogenic response * Found that MLC phosphorylation was increased equally at 3 different time points during the myogeni response

Answer 19

* Added a concentration of NA that eventually produced the same level of contraction as the myogenic response did * Found that there were lower levels of calcium in the NA experiment than the myogenic experiment -\> This suggests that while myogenic contraction involved voltage-dependent calcium entry, this might be less so the case in the NA experiment * In both experiments, MLC phosphorylation was increased at the start of the response, but in the myogenic response it stayed constant, whilei in the NA response it decreased over time * At the point where the contraction was the same, MLC phosphorylation was lower in the NA group, which suggests there may be a different mechanism at play here enabling the same level of contraction The main point is that the response to a vasoconstrictor is not sustained, while myogenic tone is sustained.

Answer 20

See previous lecture about NO, prostacyclins and EDHF

Answer 21

* At normal pressure, no. Use of various blockers of the ACh response to block NO and calcium-dependent potassium channels produced no change in myogenic tone. * However, when the luminal pressure is lowered to 5mmHg, the number of calcium events inside endothelial cells doubled * Any reduction in myogenic tone at this low pressure could be blocked using a TRPV4 antagonist, suggesting these play a role in this process * The TRPV4 channels are close to holes in the internal elastic lamina and are aligned with IK_Ca channels, which enable hyperpolarisation and relaxation of the SMCs

Answer 22

* TRPV4 * IK_Ca

Answer 23

* There is bi-directional feedback between the endothelial cells and smooth muscle cells in arteries See the flashcards from earlier lecture.

Answer 24

* Studied a preparation of endothelial cells * Addition of phenylephrine or Bay K 8644 (L-type calcium channel agonist) did not produce an increase in spontaneous activity -\> This suggests there are no calcium channels involved * Addition of ACh did lead to an increase in spontaneous activity -\> This shows this is a viable preparation and that there are muscarinic receptors present

Answer 25

* Calcium channels are present on SMCs in the arteries * There can be influx of calcium from these SMCs into the endothelial cells

Answer 26

* Application of phenylephrine (alpha-1 agonist), potassium (for depolarisation of membrane to open VGCCs) and BayK (L-type calcium channel agonist) to smooth muscle cells all led to an increase in endothelial cell calcium events * This response was blocked by nifedipine in all cases except with the high concentration phenylephrine * This suggests that VGCCs and alpha-1 receptors are in SMCs * The VGCCs lead to an increase in intracellular calcium, which enters the endothelial cells * The alpha-1 receptors lead to an increase in intracellular IP₃, which enters the endothelial cells and triggers calcium release

Answer 27

IK_Ca channels on the endothelial cells: * Addition of 3nM BayK led to slight vasoconstriction and then slight feedback vasodilation * Addition of 30nM BayK led to more pronounced vasoconstriction and then feedback vasomotion * When TRAM-34 (IK_Ca blocker) was added, there was no feedback vasodilation or vasomotion in response to BayK

Answer 28

* Luminal pressure initiates myogenic tone * It is counteracted by endothelial-cell dependent feedback, but this is only apparent at very low pressures (such as during intense vasoconstriction) * Therefore, there is a high degree of resting myogenic tone

Answer 29

* Potassium released from endothelial cells can activate (inward rectifier) potassium channels on adjacent endothelial cells. * Hence, there is spread of the hyperpolarisation. * This enables the arterioles to act as a syncytium.

Answer 30

* Measured the myogenic tone and membrane potential of an endothelial cell * Applied ACh at progressively further distances from the endothelial cell * At each distance, hyperpolarisation and vasodilation were simultaneously increased

Answer 31

* Skeletal muscle cells have K_ATP channels that are activated by low ATP and there is also general release of potassium -\> This potassium efflux can activate inward rectifier potassium channels on smooth muscle cells and endothelial cells in the arteries * ADP and adenosine are vasodilators * Some ACh from the NMJ can reach the arteries, leading to vasodilation

Answer 32

* Studied the module inflow arteriole (which supplies the skeletal muscle) and the upstream arteriole * In response to skeletal muscle contraction, both dilated * Although the module arteriole dilated by a higher percentage, since the upstream arteriole is larger, it may be responsible for most of the increased flow in the module arteriole * Use of a K_ATP channel blocker inhibited the vasodilation in the module arteriole * This suggests that K_ATPchannels are important in this vasodilation, since low ATP opens the K_ATP channels, leading to hyperpolarisation and vasodilation

Answer 33

* NO does not lead to conducted dilation * Skeletal muscle contraction does lead to conducted dilation due to released of vasoactive factors

Answer 34

No, the limit of resolution is around 300 micrometers.

Answer 35

* Usually, there is greater blood flow through endocardial vessels than epicardial vessels * However, at low pressures there is greater flow through epicardial vessels since the blood reaches these first * Adenosine can be added to remove the effect of autoregulation and find the maximal blood flow at any given perfusion pressure * The difference between the autoregulated and maximal blood flow is the coronary flow reserve

Answer 36

* The maximum increase in blood flow through the coronary arteries above the normal resting volume. * It can be tested by adding adenosine to fully dilate the vessels * Typical value = Around 5ml/min/g

Answer 37

Women's Ischaemia Syndrome Evaluation (WISE) study: * Compared women with high CRF (\>2.32) and low CRF (\<2.32) * Those with low CRF had much lower event-free survival over the 6 years of follow-up * The effect was preserved when looking at just women without coronary artery disease

Answer 38

* Luminal pressure initiates the myogenic response * Circulating and released vasoactive molecules * Physical factors

Answer 39

Initiated by: * Factors released from blood cells (e.g. ATP released from RBCs during hypoxia) * Factors released from cardiac myocytes (e.g. adenosine, K⁺) The endothelium acts as the conduit for the conducted vasodilation.

Answer 40

* Plotted a graph of vessel diameter against pressure * Showed that inhibition of nitric oxide synthase (by L-NAME) led to increased myogenic response * This suggests that NO is important for the myogenic response in coronary arteries, unlike in skeletal muscle * Block of IK_Ca and SK_Ca channels had no effect on myogenic tone * This suggests that NO plays more of a role in controlling myogenic tone and that calcium-dependent potassium channels have no effect Spontaneous endothelial cell calcium events were linked to endothelial cell-dependent feedback onto smooth muscle cells.

Answer 41

Myogenic tone: * The vasoconstriction in response to luminal pressure * Remains stable at a given pressure * Blocked by nifedipine -\> Suggests importance of calcium entry into smooth muscle cells EC-dependent dilation: * Can be induced by agonists, such as bradykinin * Involves some NO and mainly calcium-activated K⁺ channels on endothelial cells

Answer 42

* Acetylcholine drives vasodilation * Noradrenaline drives vasoconstriction -\> This is the opposite to most vascular beds

Answer 43

* Measured the myogenic tone at a specific point along the artery * Applied a vasoactive factor at progressively further distances from the point * When bradykinin was added, there were signs of conducted vasodilation * When adenosine was added, there was some slight conducted vasodilation * When an NO donor was added, there was no conducted vasodilation * When potassium ions were added, there was some conducted vasodilation * Barium (inward rectifier potassium channel blocker) blocked conducted vasodilation in all the experiments, and also reduced local vasodilation with bradykinin and the potassium ions

Answer 44

* Application of pinacidil (K_ATP channel opener) led to an increase in the diameter of an isolated mouse papillary muscle preparation * However, in a culture of human coronary microvascular endothelial cells, pinacidil did not hyperpolarise these cells * Hence, K_ATP channels are likely present in the ventricular myocytes and/or smooth muscle cells, but not the endothelial cells * Increased potassium concentration in the endothelial cell culture led to hyperpolarisation that was blocked by barium

Answer 45

* Ischaemia in ventricular tissue leads to opening of K_ATPchannels in the myocytes -\> This leads to release of potassium from the cells * This potassium activates inward rectifier potassium channels on endothelial cells, leading to hyperpolarisation * This then leads to conducted vasodilation via potassium leading to opening of inward rectifier potassium channels along the capillary

Answer 46

Around 15%

Answer 47

* The graph shows that blood flow remains constant over a range of pressures * This is a sign of myogenic tone enabling autoregulation of blood flow

Answer 48

Overall, increased pressure leads to stretch and depolarisation of the membrane and calcium entry, which drives contraction.

Answer 49

* Good evidence for TRPC6 and TRPM4 -\> These are TRP channels in the membrane * Vasoconstrictor action of an arachidonic acid derivative, 20-HETE

Answer 50

* A vasoconstrictor that contributes to myogenic tone * It is a derivative of arachidonic acid

Answer 51

(Harder, 2011): * Found that higher concentrations of 20-HETE lead to increased currents through voltage-gated calcium channels * Also found that higher concentrations of 20-HETE suppressed the outward K_Ca current via action of PKC

Answer 52

Astrocytes

Answer 53

(Welsh, 2002): * Used antisense oligodeoxynucleotides to TRPC6 to decrease TRPC6 expression * Increased the pressure incrementally from 40mmHg to 100mmHg * Compared to control, the antisense experiment showed greatly attenuated constriction -\> This is evidence of reduced myogenic tone * Cation currents were also reduced in the antisense experiment

Answer 54

* Stretch activates GPCRs, which signal via phospholipase C * This activates TRPC6, which enables cation currents that depolarise the cell

Answer 55

* TRPM4 channels are cation channels selective for sodium * They enable depolarisation of the cell, which activates VGCCs * They are regulated by cytoplasmic calcium (Gonzales, 2010): * Performed perforated patch-clamp recordings of transient inward cation currents (TICCs) in the same cell * Found that these were activated at negative potentials * The TICCs disappear following treatment with TRPM4 siRNA * 9-phenanthrol (a blocker of TRPM4) led to hyperpolarisation and dilation of the cerebral arteries * HOWEVER, 9-phenanthrol was also shown to be an activator of endothelial IK_Ca and SK_Ca channels so this makes the mechanism of action unclear

Answer 56

(Perez, 1999): * Recorded spontaneous transient outward currents (STOCs) alongside calcium sparks in the cerebral arteries. * The sparks and STOCs were at the same time. * This is evidence for calcium sparks activating STOCs through K_Ca channels. * This outward potassium current hyperpolarises the cell and suppresses further calcium influx via LTCCs.

Answer 57

* As pial arterioles penetrate the brain parenchyma, they gradually lose innervation from extracellular nerves originating from peripheral ganglia. * The vascular tone of parenchymal arterioles is chiefly regulated by signals derived from neurons and astrocytes that encase much of the surface of the arterioles. * This concept of coupling between the arterioles and surrounding cells is called the neurovascular unit.

Answer 58

* A signal of need for increased downstream blood flow can arise in levels 1 or 2 * It is conducted up to higher levels * This then results in vasodilation here

Answer 59

(Longden, 2011): * Imaged a cerebral arteriole that was linked to an astrocyte * Electrical field stimulation (EFS) of the astrocyte led to arteriole dilation * RBCs entered arteriole, showing increased flow * This dilatation was sensitive to K_Ca channel blockers and K_IR channel blockers (barium) * Thus, the suggested mechanism is that EFS (a proxy for nerve stimulation of the astrocyte) leads to an increase in astrocytic calcium, which opens K_Ca channels on the astrocyte. The increased extracellular potassium leads to opening of K_IR, which leads to hyperpolarisation and dilation. (Knot, 1996): * Demonstrated the importance of K_IR channels on smooth muscle cells * Increased extracellular K⁺ evokes hyperpolarization and relaxation of cerebral arteries * This is blocked by Ba²⁺ (K_IR blocker) * This is independent of endothelium (on smooth muscle) * Not all arteries have K_IR channels on their smooth muscle cells (e.g. mesenteric arteries) Hence, conductive vasodilation in arterioles is mediated by neurons and astrocytes releasing potassium onto the arterioles and further upstream. There is likely to be retrograde spread of the potassium upstream. However, there is also more recent evidence that glutamate release from excitatory nerves onto astrocytes and neurons can also stimulate vasodilation via release of arachidonic acid derivatives, mainly PGE₂onto the smooth muscle. They could also release NO. (Dabertrand, 2013): * Found evidence that contradicted the importance of PGE₂ in vasodilation * In parenchymal arterioles, PGE₂ produced vasoconstriction and not vasodilation

Answer 60

* Inward rectifier refers to the idea that the channels are much better at allowing potassium into the cell than out of the cell * However, most of the time, the extracellular potassium concentration means that K_IR does not show an conductance (figure C) * When the extracellular potassium is increased, the curve shifts so that the K_IRshows outwards conductance in the physiological range (figure D) * Under physiological conditions, the curve is never in a position where K_IRshows inwards conductance (hence the name is misleading) * The result of this is that the graph for observed membrane potential against [K⁺]_o is not as predicted by the Nernst equation (figure B) -\> This is because when at low [K⁺]_o, there is no conductance through the K_IR, so the membrane potential does not tend towards the equilibrium potential * What all this means is that when there is an increase in extracellular potassium (e.g. when an adjacent cell releases potassium), there is often an outward potassium current through K_IR, hyperpolarising the cell -\> In smooth muscle cells, this leads to vasodilation

Answer 61

Vasodilation: * When astrocytes release potassium, it leads to hyperpolarisation of the vascular smooth muscle cells by two mechanisms: * Activation of K_IRchannels * Activation of the Na⁺/K⁺-ATPase * This leads to closing of VGCCs and thus vasodilation Vasoconstriction: * When astrocytes release a LOT of potassium (due to enhanced neuronal activity), it leads to a situation where the potassium concentration is beyond the range where K_IR is activated * Instead, the potassium leads to depolarisation as predicted by the Nerst equation

Answer 62

* Stress-induced glucocorticoid signaling remodels neurovascular coupling by reducing cerebrovascular inwardly rectifying K+ channel function * This leads to reduced hyperpolarisation in response to extracellular potassium, so that relaxation is blunted

Answer 63

(Chen, 2014): * Showed the importance of endothelial cells in this process (showing that smooth muscle cells are not required, as they are not present in capillaries) * Optically studied the area of the cerebral cortex of rats that was activated by a somatosensory stimulus * Found that a stimulus led to an increase in blood flow to the region for 15 seconds * Used light dye damage to endothelial cells -\> This blocked the propagation of the vasodilation, suggesting that the endothelial cells are responsible for the conductive vasodilation (Longden, 2017): * Suggested that the mechanism of spread of vasodilation must be a retrograde spread of hyperpolarization from capillaries to arterioles * Application of 10mM K⁺ to capillaries led to marked vasodilation in the upstream arterioles, which was preceded by hyperpolarisation of the arterioles * This was blocked by barium (K_IR blocker) * The effect was also abolished by K_IRknockout * The results were also replicated in vivo using fluorescently-labelled RBCs as a marker of flow

Answer 64

* It has been suggested that active neurons and astrocytes produce vasodilators (mainly PGE₂) that affect pericytes wrapped around capillaries * This pericyte dilation may spread to the upstream arterioles The mechanism is shown in the diagram.

Answer 65

* Useful: V/Q matching * Harmful: HAPE

Answer 66

When there is uneven HPV, there can be very high pressures in certain capillaries, leading to oedema.

Answer 67

* A rat is prepared in a way that allows the gases flowing into the lungs and lung perfusion to be controlled * The pulmonary arterial blood pressure can also be measured

Answer 68

The blood pressure increases with each challenge, showing a degree of summation until the blood pressure reaches an optimum response.

Answer 69

(Bergofsky, 1968): * Created a setup with an isolated lung where the gases entering the lung could be varied and the blood gases could be varied using a disc oxygenator * Varied the alveolar, arterial and venous (with the flow reversed) oxygenation to see the effect on pulmonary vascular resistance * Only the alveolar hypoxia induced a large increase in pulmonary vascular resistance -\> This suggests the main sensor for HPV is in the airways * Arterial hypoxia had small effect on pulmonary vascular resistance

Answer 70

(Hakim, 1982): * Created a model where the arterial and venous pressure could be measured, as well as where the arterial and venous segments could be constricted * In this model, it is assumed that the arterial and venous segments are rigid, while the capillaries are compliant so they can buffer changes in pressure * In the control experiment, arterial occlusion led to sharp fall in arterial pressure followed by a plateauing of the arterial pressure -\> This is because the capillaries buffer the extra pressure until the capillary pressure is equal to the arterial pressure. So the plateau pressure is the capillary pressur.e * Similarly, in the control experiment, venous occlusion led to a sharp increase in venous pressure, followed by a slower increase once the capillary pressure equals the venous pressure * During hypoxia, the arterial pressure is higher, but the venous and capillary pressures remain unchanged -\> This shows that HPV is driven by arterial contraction

Answer 71

* A technique used to study pressure changes in single blood vessels * A section of artery is mounted with a force transducer (to measure force generation) and micrometer (to apply basal stretch or contraction)

Answer 72

* The vasoconstriction occurs in two phases: an initial sharp acute increase followed by a smaller gradual increase * There is a staircase effect of the phase 2 response after repeat hypoxia * The tissue-wide response does not feature a phase 1, only a phase 2 -\> The reason for this difference has not been explained

Answer 73

(Harder, 1985): * Did microelectrode recordings of a pulmonary artery smooth muscle * Hypoxia led to a gradual depolarisation and firing of action potentials in the smooth muscle * These were blocked by the verapamil, showing they were calcium-dependent (Weir, 1992): * Used patch-clamping to study the physiology of individual smooth muscle cells * Confirmed that hypoxia led to progressive depolarisation * Showed that this depolarisation was underlied by a delayed-rectifier K⁺ current that is reduced during hypoxia * However, this potassium current was only seen at high voltages (Gurney, 1997): * Used a different voltage-clamp protocol to plot a voltage-current graph for a smooth muscle cell under control and hypoxic conditions * This showed evidence of a potassium current (named I_Kn) that was suppressed by hypoxia at a range of voltages (Olschewski, 2006): * Found that the I_Kn current is likely via TASK-1 channels * This is because anandamide (TASK-1 blocker) led to a reduction in the I_Kn current * siRNA transfection confirmed this This all gives rise to the electrical model of HPV, where hypoxia inhibits potassium currents, leading to depolarisation, increased LTCC currents and vasoconstriction.

Answer 74

* Myography was used to study the hypoxic response in pulmonary smooth muscle * The phase 1 and 2 responses were still present (only slightly attenuated) in the presence of nifedipine * When the tissue was pre-depolarised using a solution of 80mmM potassium (which would inactivate the LTCCs), there was still a clear phase 1 and 2 response * This is indicative of the fact that the response might not rely on extracellular calcium influx, but internal calcium release

Answer 75

(Robertson, 1995): * Measured tension and intracellular calcium during the HPV response * Found that calcium stayed constant during the phase 2 response, but tension gradually increased * This is indicative of some form of calcium sensitisation

Answer 76

Component 3 is calcium sensitisation.

Answer 77

* Component 1 * Inhibited by cyclopiazonic acid (Calcium ATPase inhibitor) -\> Not clear why * Independent of external calcium * Component 2 * Inhibited by 8-Br-cADPR (RyR blocker) * Independent of external calcium * Component 3 * Inhibited by removal of endothelium * Dependent on external calcium

Answer 78

Component 1: * Assumed to involve SR calcium release * Thought to be due to the functioning of the calcium ATPase Component 2: * Due to cyclic ADP-ribose activation of the RyR Component 3: * Involves oxygen-sensing in the endothelium and release of a factor * This factor leads to inhibition of smooth muscle myosin phosphatase * A possible factor would be endothelin-1 In this model, the sensor of hypoxia could be mitochondrial activity, which affects production of cADPR. Another mechanism could be the production of ROS.

Answer 79

(Rounds, 1981): * Found that various poisons of the ETC led to increases in pulmonary artery pressure * This suggests that the mitochondria may affect HPV because they link to metabolism (Archer, 1993): * Proposed a ROS model, since ROS are a product of the ETC * Hence, ROS could be an indicator of oxygen levels * Chemiluminescence showed that ROS were decreased in hypoxia * Antimycin and rotenone (inhibitors of the ETC) led to decreases in ROS and an increase in pulmonary artery pressure -\> They are irreversible poisons so sensitivity to hypoxia was lost * Cyanide (also an inhibitor of the ETC) similarly led to a decrease in ROS and an increase in pulmonary artery pressure -\> Cyandine is reversible so hypoxic sensitivity returned (Chandel, 2000): * Found that myxothiazol and antimycin (both complex III inhibitors) had different effects on HPV * Myxothiazol led to ablation of HPV, while antimycin did not * This was proposed to be because they act at different points on complex III, so antimycin enables the intermediate semiquinone to be stabalised, enabling ROS production * This led to the idea that ROS production at complex III was indicator of hypoxia, underlying HPV * However, this was contradictory to the idea that ROS goes down during hypoxia, while this model suggests ROS would go up -\> This required further investigation (Desireddi, 2010): * Attempted to measure ROS production during hypoxia * Used RoGFP as an indicator -\> This is a bit of a surrogate measure since the RoGFP relies on glutathione, which is part of the antioxidant defense mechanism but not superoxide or hydrogen peroxide per se * This showed that ROS was increased in hypoxia, which could be ablated using catalase

Answer 80

* It usually has a very high affinity for oxygen, so its function would not be compromised by physiological levels of hypoxia * However, the carotid body has an unusual complex IV that has an altered oxygen affinity -\> Evidence for this in the pulmonary vasculature is currently lacking * This could be due to different isoforms * (Sommer, 2017): * Found that knockout of complex IV isoform 2 led to a lack of hypoxic response in isolated vessels * However, oxygen sensitivity did not change in the knockout mice

Answer 81

* H₂S has different effects in different vessels -\> In some it causes vasodilation and in some it causes vasoconstriction * However, the effects of hypoxia and H₂S tend to be the same in each type of vessel * (Olsen, 2010): * Showed that H₂S degradation is oxygen-dependent * Also showed that oxygen-sensitivity of H₂S degradation is unusually sensitive in the lungs * However, critics have argued that cystathionine-gamma-lyase inhibition does not affect HPV

Answer 82

(Robbins, 2008): * 8 hour exposure to hypoxia led to increase baseline pulmonary arterial pressure and also increased PAP after acute hypoxia * Iron affects the response, suggesting that HIF may play a role Chuvash polycythaemia also features increased HPV.

Answer 83

It has to take all of the blood with every cycle.

Answer 84

(West, 1964): * Utilised the setup on the previous flashcard * Used xenon¹³³ to study blood flow to different heights in the lungs * First plotted count rate against height -\> However, this is affected by the volume of lung at each height * Next plotted the same after rebreathing the gas into a bag for 2 minutes -\> This allows the xenon to equilibriate throughout the lungs, meaning that the count rate is an indicator of lung volume, not blood flow * Dividing the graphs by each other gives a plot of blood flow per unit volume at each height * This showed that blood flow decreases with height * Repeated this at a lower inflow pressure (16cmH₂O) and found that blood flow stopped at a lower height

Answer 85

Alveolar pressure can change and affect these zones -\> Positive pressure ventilation will increase the size of zones one and two.

Answer 86

* West's classic xenon experiments suggest so * However, other studies have not found this * (Prisk, 1994) found that some inhomogeneity of pulmonary blood flow appears to persist in space, in the absence of gravity

Answer 87

* Doppler echocardiography can be used to estimate pulmonary artery systolic pressure (PASP) -\> This is done by looking at the speed of tricuspid valve regurgitation during ventricular contraction * Swan-Ganz catheterization

Answer 88

(Grünig, 1997): * Found that pulmonary pressures remain relatively constant with increasing workload * During moderate exercise, distension and recruitment of pulmonary vessels means that PVR falls, and pulmonary pressures can remain relatively constant.

Answer 89

(Borst, 1956): * Created a setup including the heart and lungs of anaesthetised dogs connected to a system that allows the left atrial pressure (a proxy for pulmonary venous pressure) and pulmonary arterial pressure to be altered * Plotted the PVR against left pulmonary arterial pressure * At low pulmonary venous pressures, a small change in pulmonary arterial pressure leads to a large change in PVR * But at high pulmonary venous pressures, a large change in pulmonary arterial pressure leads to a small change in PVR * This shows how the system begins to act more like a rigid tube at high transmural pressure (i.e. it becomes less distensible)

Answer 90

Thicker, less distensible vessel walls

Answer 91

Cor pulmonale (right heart failure), which leads to: * Right ventricular hypertrophy and dilatation * Peripheral oedema

Answer 92

(Smith, 2009)

Answer 93

(Grunig, 1997): * Compared the response to 4 hours of hypoxia in control patients and those who had previously experienced HAPE * The pulmonary arterial systolic pressure increased much more in the HAPE group than the control group * This suggests HAPE may be driven by HPV In particular, uneven HPV may lead to stress failure of capillaries.

Answer 94

Inhaled beta-2 agonists (salmeterol) reduce HAPE in susceptible individuals, perhaps by enhancing alveolar fluid clearance.

Answer 95

(Chen, 1995): * Chronic hypoxia causes pulmonary hypertension and right ventricular hypertrophy in mice. * There are signs of thickening of the pulmonary arteries. This can be prevented by endothelin antagonists, or by inactivation of the hypoxia-inducible factor transcription factors. (Groves, 1987): * Volunteers were decompressed in a hypobaric chamber for 40 days to a barometric pressure (PB) of 240 Torr, equivalent to the summit of Mt. Everest. * Serial measurements of the pulmonary vascular pressure gradient were then taken at PB 760 (sea level), 347, and 282/240 Torr. * At higher altitudes, the pressure gradient increased more with cardiac output and cardiac output could be increased less * This is also evidence of increased rigidity of the pulmonary vascular system * The ‘rigidity’ of the pulmonary circulation at altitude might limit exercise capacity by placing a strain on the right ventricle

Answer 96

(Naeiji, 2010): * Found that some of the decrease in exercise capacity during acute and chronic hypoxia can be reversed using sitaxsentan (endothelin receptor antagonist)

Answer 97

(Yu, 1999): * HIF-1 deficiency in mice (HIF1α+/- ) confers protection against hypoxic pulmonary hypertension (Smith, 2006): * HIF upregulation in humans with Chuvash polycythaemia causes pulmonary hypertension (Groves, 1993): * Tibetans seem to have selected against high pulmonary pressures at altitude, possibly contributing to remarkable exercise performance * This population has been at altitude for the longest, suggesting that this response is unwanted (Petousi, 2014): * There is blunted HPV in Tibetans

Answer 98

(Dorrington, 2010): * Studied the effect of hypercapnia and hypocapnia on pulmonary arterial pressure * Hypercapnia increased the pressure compared to control, while hypocapnia decreased it This suggests that alveolar P_CO2 may be at least as important as P_O2 for the acute matching of perfusion to ventilation in the human lung.

Answer 99

TMN = Transit time of the saline bolus

Answer 100

* It can be used to identify areas of ischaemic damage (i.e. scar tissue) in the heart. * Gadolinium is used as the contrast. It is washed out of tissue very quickly, but not in the case of scar tissue. Hence, 10 minutes after a bolus of gadolinium is given, areas of scar tissue will still retain the contrast.