Neuroprotective Effect of Cardamom Oil Against Aluminum Induced Neurotoxicity in Rats

Abstract

Acetylcholinesterase (AChE) is an enzyme involved in the progression of Alzheimer's disease (AD). Cardamom oil (CO) has been reported to have acetylcholinesterase inhibitory, antioxidant and anti-anxiety effects. Hence, we studied the effect of cardamom oil in aluminum chloride induced neurotoxicity in rats. AD like symptoms were induced in Wistar rats with aluminum chloride (100 mg/kg, p.o.). Cardamom oil was administered concomitantly by oral route at doses of 100 and 200 mg/kg for 42 days. Behavioral parameters like Morris water maze, elevated plus maze, passive avoidance test and locomotor activity were evaluated on day 21 and 42. AChE activity, oxidative stress parameters, histopathological studies and immunohistochemistry studies were carried out in hippocampus and cortex. Cardamom oil treatment showed significant improvement in behavioral parameters, inhibition of AChE activity (p < 0.001) and reduction in oxidative stress in the brain. Histopathological studies of hippocampus and cortex by hematoxylin & eosin (H. & E.) and congo red stain showed inhibition of neuronal damage and amyloid β plaque formation with cardamom oil treatment. Immunohistochemistry showed, CO treatment inhibited amyloid β expression and upregulated brain-derived neurotrophic factor (BDNF). The present study showed that, cardamom oil has neuroprotective effect in aluminum chloride induced neurotoxicity linked with inhibition of AChE activity and reduction in oxidative damage. This effect of cardamom oil may be useful in management of Alzheimer's disease.

Keywords: Alzheimer's disease; acetylcholinesterase; aluminum; cardamom oil; neuroprotective; oxidative stress.

Figure 1

Effect of Cardamom oil on locomotor activity of rats using actophotometer on AlCl₃-induced neurotoxicity. Data are expressed as mean ± SEM (n = 6), ###p < 0.001 when compared with normal control, ^***P < 0.001 when compared with disease control group. AlCl₃, Aluminum chloride; SEM, Standard Error of Mean.

Figure 2

Effect of Cardamom oil on Morris water maze test in AlCl₃-induced neurotoxicity in rats. Data are expressed as mean ± SEM (n = 6), ###p < 0.001 when compared with normal control, ^**p < 0.01, ^***P < 0.001 when compared with disease control group. AlCl₃, Aluminum chloride; SEM, Standard Error of Mean.

Figure 3

Effect of Cardamom oil on transfer latency of rats using Elevated plus maze test in AlCl₃-induced neurotoxicity. Data are expressed as mean ± SEM (n = 6), ##p < 0.01, ###p < 0.001 when compared with normal control, ^*p < 0.05, ^**p < 0.01, ^***P < 0.001 when compared with disease control group. AlCl₃, Aluminum chloride; SEM, Standard Error of Mean.

Figure 4

Effect of Cardamom oil on latency of rats using Passive Avoidance test in AlCl₃-induced neurotoxicity. Data are expressed as mean ± SEM (n = 6), ###p < 0.001 when compared with normal control, ^*p < 0.05, ^**p < 0.01, ^***P < 0.001 when compared with disease control group. AlCl₃, Aluminum chloride; SEM, Standard Error of Mean.

Figure 5

Effect of Cardamom oil on acetylcholinesterase assay in (A) hippocampus and (B) cortex tissue in AlCl₃-induced neurotoxicity in rats. Data are expressed as mean ± SEM (n = 6), ##p < 0.01, ###p < 0.001 when compared with normal control, ^***p < 0.001, ^**p < 0.01, ^*p < 0.05 when compared with disease control group. AlCl₃, Aluminum chloride; SEM, Standard Error of Mean.

Figure 6

Effect of Cardamom oil on H.E. stained hippocampus and cortex tissue (400X). (a) Normal control group: Hippocampus and Cortex: showing normal histology, normal neuronal cell (small arrow) {H & E, 400X}, (b) AlCl₃ group: Hippocampus and Cortex: showing neuronal degeneration with pyknotic nuclei (small arrow), reduced layer of neuronal cell (Large arrow) {H & E, 400X}. (c) AlCl₃ + Donepezil hydrochloride (1 mg/kg): Hippocampus and Cortex: showing normal histology, focal mild neuronal cell with pyknotic nuclei (small arrow) {H & E, 400X}, (d) AlCl₃ + Cardamom oil (100 mg/kg): Hippocampus and Cortex: showing neuronal degeneration with pyknotic nuclei (small arrow), reduced layer of neuronal cell (Large arrow) {H & E, 400X}. (e) AlCl₃ + Cardamom oil (200 mg/kg): Hippocampus and Cortex: showing focal mild neuronal cell with pyknotic nuclei (small arrow) {H & E, 400X}. The magnification was 400X. H.E., Hematoxylin and eosin; AlCl₃, Aluminum chloride.

Figure 7

Effect of Cardamom oil on Congo red stained hippocampus tissue (400X). (a) Normal control: Hippocampus: showing normal histology, normal neuronal cells in (small arrow) {Congo Red, 400X}, (b) AlCl₃ group: Hippocampus: showing multifocal, moderate deposition of amyloid β plaques (arrow) {Congo Red, 400X}, (c) AlCl₃ + Donepezil hydrochloride (1 mg/kg): Hippocampus: showing normal histology, normal neuronal cells (small arrow) {Congo Red, 400X}, (d) AlCl₃ + Cardamom oil (100 mg/kg): Hippocampus: showing multifocal mild deposition of amyloid β plaques (arrow) {Congo Red, 400X}, (e) AlCl₃ + Cardamom oil (200 mg/kg): Hippocampus: showing multifocal mild deposition of amyloid β plaques (arrow) {Congo Red, 400X}, (f) Histogram of congo red staining. Data are expressed as mean ± SEM (n = 6), ###p < 0.001 when compared with normal control, ^**p < 0.01, ^*p < 0.05 when compared with disease control group. The magnification was 400 X. AlCl₃, Aluminum chloride; SEM, Standard Error of Mean.

Figure 8

Effect of Cardamom oil on Congo red stained cortex tissue (400X). (a) Normal control: Cortex: showing normal histology, normal neuronal cells in (small arrow) {Congo Red, 400X}, (b) AlCl₃ group: Cortex: showing multifocal moderate deposition of amyloid β plaques (arrow) {Congo Red, 400X}, (c) AlCl₃ + Donepezil hydrochloride (1 mg/kg): Cortex: showing multifocal mild deposition of amyloid β plaques (arrow) {Congo Red, 400X}, (d) AlCl₃ + Cardamom oil (100 mg/kg): Cortex: showing multifocal moderate deposition of amyloid β plaques (arrow) {Congo Red, 400X}, (e) AlCl₃ + Cardamom oil (200 mg/kg): Cortex: showing multifocal mild deposition of amyloid β plaques (arrow) {Congo Red, 400X} (f) Histogram of congo red staining. Data are expressed as mean ± SEM (n = 6), ### p < 0.001 when compared with normal control, ^**p < 0.01, ^*p < 0.05 when compared with disease control group. The magnification was 400X. AlCl₃, Aluminum chloride; SEM, Standard Error of Mean.

Figure 9

Effect of Cardamom oil on amyloid β expression in hippocampus tissue (400X). (a) Normal control: Hippocampus: showing normal neuronal cells without immunoreactivity (arrow){Immunostaining: Amyloid- β, 400X}, (b) AlCl₃ group: Hippocampus: showing significantly mild enhanced expression of amyloid- β, exhibited by brown coloration (arrow){Immunostaining: Amyloid- β, 400X}, (c) AlCl₃ + Donepezil hydrochloride (1 mg/kg): Hippocampus: showing normal histology, normal neuronal cells without immunoreactivity (arrow) {Immunostaining: Amyloid- β, 400X}, (d) AlCl₃ + Cardamom oil (100 mg/kg): Hippocampus: showing significantly enhanced expression of amyloid- β, exhibited by brown coloration (arrow) {Immunostaining: Amyloid- β, 400X}, (e) AlCl₃ + Cardamom oil (200 mg/kg): Hippocampus: showing normal histology, normal neuronal cells without immunoreactivity (arrow){Immunostaining: Amyloid- β, 400X}, (f) Histogram of IHC. Data are expressed as mean ± SEM (n = 6), ###p < 0.001 when compared with normal control, ^**p < 0.01, ^*p < 0.05 when compared with disease control group. The magnification was 400X. AlCl₃, Aluminum chloride; SEM, Standard Error of Mean; IHC, Immunohistochemistry.

Figure 10

Effect of cardamom oil on amyloid β expression in cortex tissue (400X). (a) Normal control: Cortex: showing normal neuronal cells without immunoreactivity (arrow){Immunostaining: Amyloid- β, 400X}, (b) AlCl₃ group: Cortex: showing significantly enhanced expression of amyloid- β, exhibited by brown coloration at cortical neurons {Immunostaining: Amyloid- β, 400X}, (c) AlCl₃ + Donepezil hydrochloride (1 mg/kg): Cortex: showing normal histology, normal neuronal cells without immunoreactivity (arrow) {Immunostaining: Amyloid- β, 400X}, (d) AlCl₃ + Cardamom oil (100 mg/kg): Cortex: showing normal histology, significantly enhanced expression of amyloid- β, exhibited by brown coloration at cortical neurons (arrow) {Immunostaining: Amyloid- β, 400X}, (e) AlCl₃ + Cardamom oil (200 mg/kg): Cortex: showing normal neuronal cells without immunoreactivity at cortex (small arrow), note weak immunoreactivity (large arrow){Immunostaining: Amyloid- β, 400X}, (f) Histogram of IHC. Data are expressed as mean ± SEM (n = 6), #p < 0.05 when compared with normal control, ^*p < 0.05 when compared with disease control group. The magnification was 400 X. AlCl₃, Aluminum chloride; SEM, Standard Error of Mean; IHC, Immunohistochemistry.

Figure 11

Effect of cardamom oil on BDNF expression in hippocampus tissue (400X). (a) Normal control: Hippocampus: showing mild enhanced expression of BDNF, exhibited by brown coloration at hippocampal neurons (arrow) {Immunostaining: BDNF, 400X}, (b) AlCl₃ group: Hippocampus: showing reduced expression of BDNF, exhibited by brown coloration at hippocampal neurons (arrow) {Immunostaining: BDNF, 400X}, (c) AlCl₃ + Donepezil hydrochloride (1 mg/kg): Hippocampus: showing moderately enhanced expression of BDNF, exhibited by brown coloration at hippocampal neurons (arrow) {Immunostaining: BDNF, 400X}, (d) AlCl₃ + Cardamom oil (100 mg/kg): Hippocampus: showing reduced expression of BDNF, exhibited by brown coloration at hippocampal neurons (arrow) {Immunostaining: BDNF, 400X}, (e) AlCl₃ + Cardamom oil (200 mg/kg): Hippocampus: showing normal histology, mild enhanced expression of BDNF, exhibited by brown coloration at hippocampal neurons (arrow) {Immunostaining: BDNF, 400X}, (f) Histogram of IHC. Data are expressed as mean ± SEM (n = 6), ##p < 0.01 when compared with normal control, ^**p < 0.01, ^*p < 0.05 when compared with disease control group. The magnification was 400X. AlCl₃, Aluminum chloride; SEM, Standard Error of Mean; BDNF, brain-derived neurotrophic factor; IHC, Immunohistochemistry.

Figure 12

Effect of cardamom oil on BDNF expression in cortex tissue (400X). (a) Normal control: Cortex: showing moderately enhanced expression of BDNF, exhibited by brown coloration at cortical neurons (arrow) {Immunostaining: BDNF, 400X}, (b) AlCl₃ group: Cortex: showing reduced expression of BDNF, exhibited by brown coloration at cortical neurons (arrow) {Immunostaining: BDNF, 400X}, (c) AlCl₃ + Donepezil hydrochloride (1 mg/kg): Cortex: showing moderately enhanced expression of BDNF, exhibited by brown coloration at cortical neurons (arrow) {Immunostaining: BDNF, 400X}, (d) AlCl₃ + Cardamom oil (100 mg/kg): Cortex: showing reduced expression of BDNF, exhibited by brown coloration at cortical neurons (arrow) {Immunostaining: BDNF, 400X}, (e) AlCl₃ + Cardamom oil (200 mg/kg): Cortex: showing mild enhanced expression of BDNF, exhibited by brown coloration at cortical neurons (arrow) {Immunostaining: BDNF, 400X,}, (f) Histogram of IHC. Data are expressed as mean ± SEM (n = 6), ##p < 0.01 when compared with normal control, ^*p < 0.05 when compared with disease control group. The magnification was 400X. AlCl₃, Aluminum chloride; SEM, Standard Error of Mean; BDNF, brain- derived neurotrophic factor; IHC, Immunohistochemistry.