Short Communication
Open Access
Reactive Oxygen Species within the Spinal Cord
Impairs Arterial Baroreflex Control of Renal
Sympathetic Nerve Activity
Maycon Igor de Oliveira Milanez, Cássia Toledo Bergamaschi, Ruy R Campos and Erika Emy Nishi*
Department of Physiology Federal, Cardiovascular Division, Universidade Federal de São Paulo, Brazil
*Corresponding author: Erika E. Nishi, Department of Physiology Federal, Cardiovascular Division, Universidade Federal de São Paulo, Brazil, Tel: +5511 5576-4848; E-mail:
@
Received: October 10, 2017; Accepted: November 03, 2017; Published: November 19, 2017
Citation: Emy Nishi E, Milanez M, (2017) Reactive Oxygen Species within the Spinal Cord Impairs Arterial Baroreflex Control of Renal Sympathetic Nerve Activity. SOJ Neurol 4(1), 1-4. DOI: 10.15226/2374-6858/4/1/00134
Abstract
Clinical and experimental studies have shown that sustained
increase in renal sympathetic nerve activity (rSNA) contributes to
hypertension. Previous studies by our group showed that antioxidant
treatments improve arterial baroreceptor reflex, which is a powerful
beat-to-beat negative feedback control of arterial blood pressure
(BP). We hypothesized that reactive oxygen species (ROS) contribute
to arterial baroreflex control of rSNA by acting on sympathetic
preganglionic neurons in the spinal cord. We then performed
the intrathecal (i.t.) administration of tempol (4-hydroxy-2, 2, 6,
6-tetramethylpiperidine-N-oxyl) (5nmol), a superoxide dismutase
mimetic. We evaluated the cardiovascular effects, basal and reflex
rSNA evoked by i.t. injection of tempol. No significant changes in
basal levels of heart rate (HR), mean arterial pressure (MAP) and
rSNA were found. However, tempol significantly increased renal reflex
sympathoinhibitory responses (pre tempol: -0.61 ± 0.15; post tempol:
-1.55 ± 0.14* spikes/s/mmHg). Thus, the results suggest that ROS
exert a tonic inhibitory influence on the activity of spinal neurons
with barosensitive properties, and it seems to be a preferential
influence on fibers involved in reflex sympathoinhibitory responses.
Our study raises the possibility that ROS in the spinal cord mediate
part of the baroreflex dysfunction in cardiovascular diseases, such
as hypertension and heart failure, which is detrimental to patient
outcome.
Keywords: spinal cord; renal sympathetic activity; arterial baroreflex; reactive oxygen species;
Keywords: spinal cord; renal sympathetic activity; arterial baroreflex; reactive oxygen species;
Introduction
Spinal Reactive Oxygen Species Contributes to Renal
Baroreflex Control
The arterial baroreceptor reflex is a powerful beat-tobeat
negative feedback control of arterial blood pressure.
Baroreceptors are mechanoceptors located in the aortic arch
and at the bifurcation of the carotid arteries, being sensitive to
the stretching of such vessels when arterial pressure is changed.
Signals from these structures are centrally integrated, evoking
changes in the pattern of sympathetic vasomotor activity
controlling BP (Vasquez et al., 1997; Campos et al., 2001). In order
to clarify the physiological aspects of the respective reflex, the brain circuitry involved in the modulation of this mechanism has
been extensively studied, as well as its main neurotransmitters
(Gordon, 1987; Lawrence e Jarrott, 1994; Tan et al., 2007).
Moreover, the existence of spinal neurons with barosensitive
properties has been described, bringing to light the important
role of the spinal cord in the reflex control of BP (Mccall et al.,
1977; Campos et al., 2017). However, the spinal cord mechanisms
involved in this transmission need to be clarified.
Previous studies point to the involvement of reactive oxygen species (ROS) in cardiovascular diseases, and it seems to be an important molecular target to attenuate the systemic effects evoked by such disorders (Han et al., 2007; Nishi et al., 2013). Furthermore, evidence indicates that ROS, when in excess on the system, have the ability to influence baroreflex by attenuating the mechanism sensitivity, suggesting that the existence of oxidative balance is crucial for normal reflex activity (Li et al., 1996; Campos et al., 2015). In fact, our group showed that treatment with Vitamin C for seven consecutive days significantly increased baroreceptor sensitivity not only in animals with renovascular hypertension, but also in normotensive animals (Nishi et al., 2010). Corroborating with these findings, another study showed that the reduction of ROS in the rostral ventrolateral medulla (RVLM) was able to increase the sensitivity of the baroreflex response in spontaneously hypertensive stroke prone rats (Ogawa et al., 2012). For more details about the neural control of BP we recommended the review wrote by Thomas (2011) (Thomas, 2011). Little is known about the physiological role played by spinal ROS on basal and reflex modulation of rSNA. Therefore, the present study aimed to evaluate the cardiovascular responses and the rSNA evoked by the i.t. administration of tempol, a superoxide dismutase (SOD) mimetic. In addition, we assessed the arterial baroreceptor reflex sensitivity to the renal territory after spinal administration of tempol, investigating a possible participation of the spinal ROS in the reflex control of the sympathetic vasomotor activity to the renal bed.
Previous studies point to the involvement of reactive oxygen species (ROS) in cardiovascular diseases, and it seems to be an important molecular target to attenuate the systemic effects evoked by such disorders (Han et al., 2007; Nishi et al., 2013). Furthermore, evidence indicates that ROS, when in excess on the system, have the ability to influence baroreflex by attenuating the mechanism sensitivity, suggesting that the existence of oxidative balance is crucial for normal reflex activity (Li et al., 1996; Campos et al., 2015). In fact, our group showed that treatment with Vitamin C for seven consecutive days significantly increased baroreceptor sensitivity not only in animals with renovascular hypertension, but also in normotensive animals (Nishi et al., 2010). Corroborating with these findings, another study showed that the reduction of ROS in the rostral ventrolateral medulla (RVLM) was able to increase the sensitivity of the baroreflex response in spontaneously hypertensive stroke prone rats (Ogawa et al., 2012). For more details about the neural control of BP we recommended the review wrote by Thomas (2011) (Thomas, 2011). Little is known about the physiological role played by spinal ROS on basal and reflex modulation of rSNA. Therefore, the present study aimed to evaluate the cardiovascular responses and the rSNA evoked by the i.t. administration of tempol, a superoxide dismutase (SOD) mimetic. In addition, we assessed the arterial baroreceptor reflex sensitivity to the renal territory after spinal administration of tempol, investigating a possible participation of the spinal ROS in the reflex control of the sympathetic vasomotor activity to the renal bed.
Methods
Animals
The experimental protocols employed in this study were
performed under guidelines recommended by the National
Institutes of Health and approved by the Ethics in Research
Committee of the Escola Paulista de Medicina - Universidade
Federal de São Paulo (process No.8724270715/15). Male Wistar
rats (n=4) (300–350g) were housed in group cages, given access to
rat chow and water ad libitum, and maintained in a temperaturecontrolled
environment (23 °C) with a 12-hour light/dark cycle.
Recording of mean arterial pressure and heart rate
Rats under ketamine (80-100 mg/kg, ip) and xylazine (10
mg/kg, ip) anesthesia had the femoral vein catheterized for
intravenous injection of drugs and the femoral artery catheterized
for direct recording of arterial pressure. After surgical recovery
(approximately 24 hours), baseline pulsatile BP, MAP and HR
were recorded in conscious rats (PowerLab –ADInstruments,
Australia). Average values of basal MAP and HR were obtained by
continuous recording for 10 minutes.
Intrathecal Injection
In urethane-anesthetized rats (1.2 to 1.4 g/kg) (SigmaAldrich,
St. Louis, MO), the spinal subarachnoid space was cannulated for
the i.t administration of drugs. The atlanto-occipital membrane
was exposed by removing the overlying neck musculature
through a midline dorsal incision. The head of the rat was mildly
depressed and the membrane was carefully slit. A polyethylene
(PE-10) catheter filled with tempol (5nmol in 2μL - SigmaAldrich,
St. Louis, MO) dissolved in saline was advanced caudally in the
spinal sub-arachnoid space until T11-12 segment of spinal cord,
where the drug was injected. The drug effects were analyzed over
10 minutes; the validity of this method for correct positioning of
the tip of the catheter was confirmed in post-mortem analysis.
Analysis of renal sympathetic nerve activity (rSNA)
and the arterial baroreflex control of rSNA in rethaneanesthetized
rats
The left renal nerve was retroperitoneally exposed and placed
on bipolar silver electrodes. The signal from the renal nerve was
displayed on an oscilloscope TDS 220 (Tektronix, Beaverton, OR).
The nerve activity was amplified (gain 20 K Neurolog; Digitimer,
Welwyn Garden City, Herts, UK), filtered by a band-pass filter (100–
1000 Hz), and collected for display and subsequent analysis using
a PowerLab data acquisition system (ADInstruments, Sydney,
Australia). At the end of the experiments, the background noise
level was determined by hexamethonium bromide administration
(30mg/kg, intravenously) (SigmaAldrich, St. Louis, MO). The
neural activity was analyzed using the appropriate software spike
histogram and was expressed as spikes/second (ADInstruments).
For analysis of the arterial baroreflex control of rSNA, bolus injections (0.1 ml) of phenylephrine (10 μg, iv) (SigmaAldrich, St. Louis, MO) and sodium nitroprusside (20 μg, iv) (SigmaAldrich, St. Louis, MO) were administered pre and post i.t. injection of tempol. Values of matching MAP variations (ΔMAP from 5 to 40 mm Hg) with reflex rSNA (ΔrSNA) responses were plotted separately for each vasoactive drug to measure baroreceptor function, and their gain (spikes/s/mmHg) were compared to evaluate changes in arterial baroreflex sensitivity.
For analysis of the arterial baroreflex control of rSNA, bolus injections (0.1 ml) of phenylephrine (10 μg, iv) (SigmaAldrich, St. Louis, MO) and sodium nitroprusside (20 μg, iv) (SigmaAldrich, St. Louis, MO) were administered pre and post i.t. injection of tempol. Values of matching MAP variations (ΔMAP from 5 to 40 mm Hg) with reflex rSNA (ΔrSNA) responses were plotted separately for each vasoactive drug to measure baroreceptor function, and their gain (spikes/s/mmHg) were compared to evaluate changes in arterial baroreflex sensitivity.
Data Analysis
The results are presented as the mean ± SD. The data were
evaluated by paired Student’s t-test. The level of statistical
significance was defined as P < 0.05.
Results and Discussion
The respective baseline values (HR, MAP and rSNA) of the
animals are presented in the figure legends.
Effects of intrathecal administration of tempol
I.t. administration of tempol unchanged basal levels of
HR, MAP and rSNA (Figure 1). However, tempol significantly
increased renal reflex sympathoinhibitory responses evoked by
intravenous injection of phenylephrine (pre tempol: -0.61 ± 0.15;
post tempol: -1.55 ± 0.14* spikes/s/mmHg) (Figure 2), whereas
no change occurred in sympathoexcitatory responses elicited by
intravenous injection of sodium nitroprusside (pre tempol: 0.47
± 0.07; post tempol: 0.37 ± 0.05* spikes/s/mmHg).
Figure 1: Responses evoked by intrathecal administration of tempol
(5nmol) on heart rate (HR, bpm) (A), mean arterial pressure (MAP,
mmHg) (B) and renal sympathetic nerve activity (rSNA, spikes/s) (C)
presented as changes from baseline. Baseline values of HR, MAP and
rSNA are: 474 ± 21 bpm, 100 ± 6 mmHg, 111 ± 9 spikes/s. Intrathecal
administration of the antioxidant did not trigger any robust change in
the HR, MAP and rSNA.
Figure 2: Analysis of the renal baroreceptor reflex. The figure shows
that the intrathecal application of tempol triggers an increase in the gain
of the sympathoinhibitory reflex response caused by phenylephrine. No
significant change was observed regarding the sympathoexcitatory reflex
responses. *p < 0.05 (paired Student’s t-test). Mean Arterial Pressure
(MAP); Renal Sympathetic Nerve Activity (rSNA).
The results indicate a physiological role played by ROSs in the
spinal cord regarding the control of rSNA. Although apparently
not involved in the basal modulation of rSNA under physiological
conditions, ROS appear to exert a tonic inhibitory influence on
sympathetic preganglionic neurons with barosensitive properties
in the spinal cord, apparently with a preferential influence on
unmyelinated fibers involved in reflex sympathoinhibitory
responses (Figure 3). Such findings suggest that, in addition to
oxidative stress in specific brain areas involved in the control of
sympathetic vasomotor tone (Ogawa et al., 2012; Campos et al.,
2015), the spinal oxidative imbalance may also be contributing
to the baroreceptor dysfunction seen in several cardiovascular
diseases (Sawyer et al., 2002; Oliveira-Sales et al., 2008).
Figure 3: Representative image of the effect evoked by intrathecal administration of tempol on the reflex sympathoinhibition to the renal territory
caused by the intravenous injection of phenylephrine. After the spinal application of the antioxidant, a more intense inhibitory effect is observed triggered
by the pressor substance. These findings suggest that ROS exert an inhibitory effect on spinal neurons with barosensitive properties, especially
those involved in reflex sympathoinhibition responses. Pulsatile Arterial Pressure (PAP); Renal Sympathetic Nerve Activity (rSNA).
Moreover, the results of the present study corroborate with
recent findings obtained in our laboratory. We have shown that
i.t. administration of losartan, an angiotensin II (Ang II) AT1
receptor antagonist, triggers a selective increase in sensitivity for
renal sympathoinhibitory reflex responses in the renovascular
hypertension model (Campos et al., 2017). The production
of Ang II-mediated ROS is widely described in the literature
(Seshiah et al., 2002; Han et al., 2007), therefore, it is reasonable
to hypothesize that the baroreceptor dysfunction found in the
respective hypertension model is evoked, in part, by a prooxidative
spinal angiotensinergic action. However, further studies
are needed to clarify the origin of spinal ROS, its topographic
location, as well as the precursor agents involved in its synthesis.
Therefore, the fact that ROS contribute to arterial baroreflex control of rSNA by acting on sympathetic preganglionic neurons in the spinal cord under physiological condition raises the possibility that spinal ROS mediate part of the baroreflex dysfunction that is detrimental to the outcome of hypertensive and heart failure patients.
Therefore, the fact that ROS contribute to arterial baroreflex control of rSNA by acting on sympathetic preganglionic neurons in the spinal cord under physiological condition raises the possibility that spinal ROS mediate part of the baroreflex dysfunction that is detrimental to the outcome of hypertensive and heart failure patients.
Financial Support
FAPESP (2013/13332-1); CAPES (0523/2014) and CNPq
(472613/2013-8). C.T.B. and R.R.C. are recipients of CNPq
fellowships. E.E.N. was the recipient of FAPESP (2013/23741-6)
and CAPES-PNPD Postdoctoral Fellowships.
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