A single blinded non-crossover investigation into the cardiovascular effects of the ?-adrenergic receptor antagonist pindolol on heart rate and systolic blood pressure during exercise on a cohort of second year medical students Introduction:During exercise, our body has higher requirements for oxygen and energy. The cardiovascular system adjusts to these demands by increasing the heart rate (HR) and stroke volume (SV)[1]. These changes are brought about by increased activity of the noradrenergic sympathetic nervous system (SNS) which lead to a higher HR and blood pressure (BP)[2].  ?-adrenergic antagonist drugs work to reduce these effects by blocking ?-receptors and hence preventing sympathetic stimulation via catecholamines. ?-blockers are used as therapy for hypertension[3], as well as treatment for angina and atrial fibrillation.This practical aims to identify the actions of maximal ?-blockade -adrenoceptor antagonists (pindolol) on cardiovascular and respiratory systems. This is achieved by observing parameters such as HR and BP. Results recorded will be compared to those produced by the control group who took the placebo drug.The hypothesis –  that the ?-blocker will cause HR to be lower in the participants that have taken the drug in comparison to the placebo, before and during exercise. This is based upon the findings of Giulia Sandrone,Andrea Mortara,Daniela Torzillo,Maria Teresa La Rovere,Alberto Malliani,Federico Lombardi[4].It is also expected that ?-blockers will cause participants who have taken pindolol to have a lower SBP before and after exercise in comparison to those who have taken the placebo based on the finding of Sripal Bangalore,David Wild,Sanobar Parkar,Marrick Kukin,Franz Messerli[5].Methodology:In the study, a blinded, non-crossover trial was used to measure to measure the effects of 5mg pindolol on ?-receptor blockade. A total of 25 trials were carried out on 51 healthy participants.To begin, the participants sat on the Monark-Ergomadic-Ergometer and baseline parameters were measured. HR was taken using a pulse oximeter whilst BP was measured by positioning a digital sphygmomanometer on the wrist (Omron RS2 BP monitor).Participants cycled for 10 minutes at 2kp resistance and 70rpm. The participant’s heart rate was measured at every minute during exercise. BP was also recorded within one minute of exercise completion. Each participant was then given either drug A (placebo) or drug B (pindolol). The participants then had 30 minutes of rest time. Then, baseline HR and BP were taken Exercise protocol was repeated ending with measuring post-exercise BP.The raw data for HR (paired parametric data) and BP (paired parametric data) were analysed using a Kolmogorov-Smirnoff test finding data to be normally distributed therefore t-tests were eligible for use in their analysis (Microsoft Excel data analysis toolpak). HR and SBP were analysed using t-tests. Standard error of mean (SEM) was calculated to demonstrate the usefulness of mean values in approximation of the population mean – represented using error bars.Systolic blood pressure (SBP) was used for the analysis as opposed to diastolic blood pressure (DBP) since SBP is considered to be more clinically useful. Also, DBP varies during exercise which makes it’s use for indication of BP inaccurate. [7][8]Results Table 1: Table comparing the heart rate changes before and after taking the placebo and before and after taking pindolol. HR0 is the heart rate measurement before exercise, HR1-9 are the measurements during exercise and HR10 is the measurement after exercise. HR0 = heart rate at 0 minutes, HR1 = heart rate at 1 minute… etc.Table 1 shows that during exercise, the HR increased before and after both drugs. The placebo significantly increased HR at minute 0. Pindolol significantly decreased HR at minutes 5-10. (6/11 data points).Figure 1: Graph showing Heart Rate increase during exercise before and after taking the placebo. Error bars represent ±SEM. (* = p<0.05)*Figure 1 shows exercise significantly increased HR both pre and post-drug from 78.20?2.42 and 85.84?4.04 bpm to 113.76?5.72 and 116.92?5.50 bpm respectively. From minute 0 to 1, the HR values both increase greatly. Between minutes 2-5 the HR increases both pre-drug and post-drug. Between minutes 6 and 7 there is very little difference between both mean values. Following this, from minutes 8-10, both mean values continue to increase. Although the placebo caused a significant increase at minutes 0 and 5, it had no significant effect on HR for most results.Figure 2: Graph showing Heart Rate increase during exercise before and after taking pindolol. Error bars represent ±SEM. (* = p<0.05)*******Figure 2 demonstrates that overall, exercise significantly increased HR. Prior to administration of Pindolol, the HR increased from 80.81?2.80 to 149.19?5.45bpm whilst after Pindolol was taken, the mean HR increased from 88.69?2.89 to 129.50?5.95. Furthermore, the difference between these results also increases, thus demonstrating Pindolol resulted in a significant decrease in overall HR. However, Pindolol was insignificant between minutes 1-2.Generally, the placebo had no significant influence on HR whilst Pindolol caused a significant decrease in HR.Table 2: Table comparing the blood pressure changes before and after taking the placebo and before and after taking pindolol. HR0 is the heart rate measurement before exercise, HR1-9 are the measurements during exercise and HR10 is the measurement after exercise. HR0 = heart rate at 0 minutes, HR1 = heart rate at 1 minute… etc.Table 2 shows both the effects of drugs and the effects of exercise on BP.  It shows that there was a significant increase between SBP pre-exercise and post-exercise before both drug A and drug B. however, there was no significant increase in SBP after placebo and/or pindolol was taken. Both the placebo and pindolol significantly decreased SBP pre-exercise but had no significant effect post-exercise.Figure 3: Bar chart showing the effect of the placebo and pindolol on systolic blood pressure (SBP) before and after exercise. Error bars represent ±SEM. (* = p<0.05)**Figure 3 shows that exercise caused no significant increase in SBP both pre-drug and post-drug in groups A and B. Figure 3 also shows there is no significant decrease between SBP pre-drug and post drug in both groups A and B.Both the placebo and pindolol had no significant effect on SBP pre-exercise and post-exercise.Discussion and ConclusionOverall, analysis found pindolol to significantly decrease HR during exercise compared to the group without ?-blocker which agrees with the hypothesis. However, pindolol had no significant effect on SBP before and after exercise thus rejecting the hypothesis.The hypothesis states that ?-blockers reduce SBP, a prediction made based on their effect on ?1-adrenoceptors however pindolol had no significant effect, disagreeing with prior literatures[11][12] (table 2). During exercise, HR and BP both increase to meet raised physiological oxygen and nutrient demands[9]. Whilst exercising, arterioles supplying working muscles vasodilate which leads to total peripheral resistance (TPR) increasing. Thus, cardiac output (CO) increases to raise BP to meet the increasing oxygen demands. The rise in CO exceeds the fall in TPR therefore BP somewhat increases with exercise[10].The SNS and renin-angiotensin system (RAS) have major roles in BP regulation[13]. ?-blockers work on the ?1-adrenoceptors on the juxtaglomerular apparatus of the kidneys causing a suppression of renin release which subsequently reduces production of angiotensin II – a powerful vasoconstrictor and a precursor of aldosterone.Angiotensin II works by upregulating cAMP in nerves of the SNS, resulting in influx of calcium ions, raising secretion of noradrenaline (NA). NA encourages vasoconstriction using alpha-adrenoceptors and increases BP. Aldosterone is a hormone which acts to raise BP by increasing the blood volume through increased water reabsorption.[14] Though pindolol works to suppress renin secretion, this investigation demonstrated a rise in SBP, possible accounted for by the short break of 30-minutes left between pre-drug and post-drug exercise. Renin has a half-life of up to 90 minutes[15] therefore circulating renin from pre-drug exercise mightn’t have been metabolised.  Consequently, remaining renin from pre-drug exercise might have exaggerated post-drug exercise SBP readings.Pindolol is a fairly lipophilic drug and hence is able to move across the blood-brain barrier.[17] Hence, pindolol can cause blockade of central ?-receptors – decreasing sympathetic domination.[18] Pindolol also blocks peripheral pre-synaptic ?1-adrenoceptors. Together these mechanisms should lower the amount of NA yet the partially-antagonistic activity of pindolol before exercise might have caused increase in NA levels, withdrawing it’s antagonistic effects on the central nervous system. Resultingly, SBP did not decrease (table 2).Moreover, pindolol acts to antagonise ?2-adrenoceptors of the skeletal muscle resulting in vasoconstriction [16] – a possible explanation for the raised SBP (figure 3). However, the effects described where overpowered by alpha-adrenoceptor-mediated vasoconstriction of other tissues and hence were negligible.Pindolol was found to partially-significantly decrease the HR during exercise in comparison to the placebo group, agreeing with the literature.[4,19,20] The sinoatrial node (SAN) is the pacemaker of the heart. Depolarisation of the SAN creates an action potential (AP), depolarising the heart’s muscle cells. Pacemaker cells have F-channels which open and inwardly conduct upon membrane polarisation, causing depolarisation of the Na+ current. Upon membrane potential reaching it’s threshold (-40mV), an AP is generated. This is caused by entry of calcium ions through L-type calcium channels. Consequently, the AP is conducted by proximal cardiac cells.In human cardiomyocytes, 70-80% of the ?-adrenoceptors are made up ?1-adrenoceptors. Exercise increases sympathetic tone, causing increased binding of NA to the G-protein coupled ?1-adrenoceptors. Activation of ?1-adrenoceptors results in stimulation of the heterotrimeric Gs protein. The ?s related to GDP dissociates, causing the replacement of GDP with GTP. The complex of the ?s and GTP bins to adenylate cyclase, converting ATP to cAMP which binds to the F-type channels and raise pacemaker permeability. Thus, sodium ions move into the cells faster, decreasing time taken for threshold to be reached so HR increases.Pindolol results in negative chronotropism via decreased catecholaminergic stimulation of ?1-adrenoceptors. Therefore post-drug HR should be lower than pre-drug HR during exercise. This was untrue during minutes 1-4 (figure 2), disagreeing with the hypothesis that the ?-blocker will cause HR to be lower in the participants that have taken pindolol. This may be due to pindolol failing to extend to levels essential in reaching maximal ?-blockade. Numerous prior studies[21,22] have suggested maximal ?-blockade occurs after two-hours – far longer than the 30 minutes this study allowed.A positive ionotropic effect is also induced by stimulation of ?1-adrenoceptors. [23] Elevated cAMP levels upregulates protein kinase A (PKA), which phosphorylates the L-type calcium channel – allowing increased calcium entry. PKA also increased activity of calcium release channels via upregulation of ryanodine receptors. These mechanisms elevate intracellular calcium, allowing increased binding of calcium-ions to troponin I, promoting troponin displacement. This allows increased binding of myosin heads, which raises contractility and CO.[23]  CO is reduced by pindolol’s negative ionotropism. In aims of maintaining BP, baroreceptor firing to the rostral-ventrolateral-medulla decreases causing sympathetic tome to increase, allowing increased binding of NA to ?1-adrenoceptors which results in vasoconstriction. This allows maintenance of BP during ?-blockade. This, alongside minimal time for pindolol to induce maximal ?-blockade may explain failure of BP to decrease (figure 3).A limitation of this study is the small sample size used (25 drug A and 26 for drub B). Smaller sample sizes result in less reliable results. [24]  Another limitation in this study was the difficulty in standardising cycling speed and resistance – this may explain the variable HR measurements. This difficulty may be due to varying lifestyle factors such as smoking and caffeine intake. To combat this, the selection criteria should have been more specific.Another major limitation in this study is the small, 5mg dose of pindolol administered. NICE suggests that a daily 15mg dose is an appropriate dose within its therapeutic window. [25] this may explain pindolol’s failure to significantly reduce SBP(figure 3).Pindolol is not the first-line-treatment for hypertension.[26]  A Pindolol caused a significant reduction in HR during exercise, agreeing with previous literature. Pindolol did not cause any significant difference in SBP before or after exercise, conflicting with literature. This is likely to be due to the limitations discussed above. References – 20271 –  Laughlin M. Cardiovascular response to exercise. 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