Objective:

2

To evaluate the therapeutic effects of Kuanxiong Aerosol (KXA) on ischemic stroke with reperfusion and elucidate the underlying pharmacological mechanisms.

Methods:

2

In vivo

pharmacological effects on ischemic stroke with reperfusion was evaluated using the transient middle cerebral artery occlusion (t-MCAO) mice model. To evaluate short-term outcome

30 mice were randomly divided into vehicle group (

n

=15) and KXA group(

n

=15). Mice in

KXA and vehicle groups received 69 mg KXA and vehicle for 1 day

respectively. To evaluate long-term outcome

35 mice were randomly divided into sham group (

n

=5)

vehicle group (

n

=15)

and KXA group (

n

=15). Mice in KXA and vehicle groups received 69 mg KXA and vehicle for 7 days

respectively. Pathological changes in the brain were observed by 2

3

5-triphenyltetrazolium chloride or Nissl stainings

and behavioral assessments

including the Modified Neurologic Severity Score

Bederson score

rotarod test

and adhesive removal test were conducted. The penetration ability of KXA and KX (KXA without propellants) through the blood–brain barrier was assessed both

in vitro

using a transwell model and

in vivo

. Furthermore

in vitro

effects of KX (5

10

and 20 μL/L) on oxygen and glucose deprivation/re-oxygenation (OGD/R)-induced injury

transient receptor potential vanilloid type 1 (TRPV1) modulation

calcium influx

and mitochondrial function were explored through Western blot

CCK-8 assay

JC-1 staining

calcium imaging

adenosine triphosphate (ATP) and antioxidant measurements.

Results:

2

In

in vivo

experiments

KXA reduced brain infarct volume and neuron loss in t-MCAO mice. Behavioral assessments showed marked improvement in the neurological deficit of t-MCAO mice with KXA treatment (

P

<

0.05 or

P

<

0.01). Additionally

in vitro

findings indicated that KX ameliorated OGD/R-induced injury through TRPV1 channel modulation. KX increased cell viability in OGD/R-treated SH-SY5Y cells and prevented OGD/R-induced calcium overload by downregulating TRPV1 expression and constraining calcium influx through TRPV1 (

P

<

0.05 or

P

<

0.01). Furthermore

KXA maintained the membrane potential and function of mitochondria in OGD/R-treated SH-SY5Y cells.

Conclusions:

2

KXA could attenuate ischemic stroke injury through TRPV1 channel modulation

indicating its potential as a promising therapeutic option for stroke in clinical practice.