Medullary Endocannabinoids Contribute to the Differential Resting Baroreflex Sensitivity in Rats with Altered Brain Renin-Angiotensin System Expression

Abstract

CB1 cannabinoid receptors are expressed on vagal afferent fibers and neurons within the solitary tract nucleus (NTS), providing anatomical evidence for their role in arterial baroreflex modulation. To better understand the relationship between the brain renin-angiotensin system (RAS) and endocannabinoid expression within the NTS, we measured dorsal medullary endocannabinoid tissue content and the effects of CB1 receptor blockade at this brain site on cardiac baroreflex sensitivity (BRS) in ASrAOGEN rats with low glial angiotensinogen, normal Sprague-Dawley rats and (mRen2)27 rats with upregulated brain RAS expression. Mass spectrometry revealed higher levels of the endocannabinoid 2-arachidonoylglycerol in (mRen2)27 compared to ASrAOGEN rats (2.70 ± 0.28 vs. 1.17 ± 0.09 ng/mg tissue; P < 0.01), while Sprague-Dawley rats had intermediate content (1.85 ± 0.27 ng/mg tissue). Microinjection of the CB1receptor antagonist SR141716A (36 pmol) into the NTS did not change cardiac BRS in anesthetized Sprague-Dawley rats (1.04 ± 0.05 ms/mmHg baseline vs. 1.17 ± 0.11 ms/mmHg after 10 min). However, SR141716A in (mRen2)27 rats dose-dependently improved BRS in this strain: 0.36 pmol of SR141716A increased BRS from 0.43 ± 0.03 to 0.71 ± 0.04 ms/mmHg (P < 0.001), and 36 pmol of SR141716A increased BRS from 0.47 ± 0.02 to 0.94 ± 0.10 ms/mmHg (P < 0.01). In contrast, 0.36 pmol (1.50 ± 0.12 vs. 0.86 ± 0.08 ms/mmHg; P < 0.05) and 36 pmol (1.38 ± 0.16 vs. 0.46 ± 0.003 ms/mmHg; P < 0.01) of SR141716A significantly reduced BRS in ASrAOGEN rats. These observations reveal differential dose-related effects of the brain endocannabinoid system that influence cardiovagal BRS in animals with genetic alterations in the brain RAS.

Alterations in the Medullary Endocannabinoid System Contribute to Age-related Impairment of Baroreflex Sensitivity

“…The BRS values obtained after SR141716A microinjection are similar to published values of baseline BRS in young SD rats (13, 27), indicating that NTS CB 1 receptor blockade normalized impaired BRS in these older animals. These results are consistent with previous observations that CB 1 blockade in the NTS of younger SD rats had no effect on BRS but dose-dependently improved BRS in (mRen2)27 rats (11). …”

“…Increased dorsal medullary 2-AG content also occurs in young adult transgenic (mRen2)27 hypertensive rats (11), which feature downregulated NTS Ang-(1–7) tone associated with markedly impaired BRS for control of HR (12) compared to young SD rats and transgenic Ang-deficient ASrAOGEN rats with enhanced baseline BRS (25). Therefore, the dorsal medulla of aged SD rats represents the second in vivo setting in which an increased Ang II-to-Ang-(1–7) RAS imbalance is associated with impaired BRS and increased levels of 2-AG.…”

“…Reflex testing was repeated 60 minutes after bilateral microinjection of the CB 1 receptor-selective antagonist SR141716A (obtained from the National Institute on Drug Abuse) into the NTS, and was completed within 15 minutes, with each rat serving as its own control. The dose and timecourse of the study were chosen based on previous experience with SR141716A microinjections in our laboratory (11) in which the effect of the drug on BRS was immediate and sustained over 60 minutes without changing resting MAP or HR. Cardiac chemosensitive vagal fiber activation was assessed at the time of BRS testing by measuring depressor and bradycardic responses to intravenous phenylbiguanide (10 µg/kg in 50 nL 0.9% NaCl) after a 5-second bolus injection.…”

“…Ang II exerts a widespread influence over the sympathetic nervous system and impairs BRS during aging (8), and may stimulate production of endocannabinoids through actions at Ang II type 1 (AT 1 ) receptors (9, 10). We previously demonstrated that blockade of CB 1 receptors within the solitary tract nucleus (NTS) of the dorsomedial medulla, the primary site for termination of baroreceptor afferent neurons, improves the blunted BRS for control of HR in young adult (mRen2)27 rats with Ang II-dependent hypertension, with no effect in age-matched normotensive Sprague-Dawley (SD) rats with normal baroreflex function (11). The impaired BRS in (mRen2)27 rats is also associated with loss of Ang-(1–7) facilitation of the BRS (12), resembling the brain RAS profile of aged SD rats (13).…”

“…We also employ mass spectrometry to measure levels of dorsal medullary endocannabinoids, and RT-qPCR to measure mRNA levels of cannabinoid receptors (CB 1 and CB 2 ), and cannabinoid receptor interacting protein (CRIP)1a (14) in the NTS of younger and older SD rats. Based on evidence for decreased NTS Ang-(1–7) tone in aging (13) and the association between augmented Ang II and endocannabinoid tone in hypertensive rats with impaired BRS (11), we hypothesized a similar trend for endocannabinoid levels in aged rats. The present study provides functional and biochemical evidence that components of the endocannabinoid system within the NTS change during normal aging and that enhanced CB 1 receptor actions contribute to declining baroreflex function with age.…”