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Journal of Chinese Integrative Medicine ›› 2011, Vol. 9 ›› Issue (11): 1254-1263.doi: 10.3736/jcim20111115

• Original Experimental Research • Previous Articles     Next Articles

Amelioration of immobilization stress-induced biochemical and behavioral alterations and mitochondrial dysfunction by naringin in mice: Possible mechanism of nitric oxide modulation

Gollapalle L. Viswanatha1(), Hanumanthappa Shylaja2, K. Sadashiva Sandeep Rao3, Yathiraj Ashwini4, V. Ramaiah Santhosh Kumar5, C. Gangadharaiah Mohan6, Venkate Gowda Sunil7, M. Venkateshappa Sarvesh Kumar8, Subbanna Rajesh1#br#   

  1. 1. Department of Pharmacology, People's Education Society College of Pharmacy, Bangalore 560050, India
    2. Department of Pharmacognosy, People's Education Society College of Pharmacy, Bangalore 560050, India
    3. Biomedical Research Centre, Sheffield Hallam University, Sheffield S11 W, UK
    4. Department of Biochemistry, Kasturba Medical College, Mangalore 575001, India
    5. Department of Pharmacology, Al-Ameen College of Pharmacy, Bangalore 560027, India
    6. Department of Pharmacognosy, Government College of Pharmacy, Bangalore 560027, India
    7. Department of Biotechnology, Oxford College of Sciences, Bangalore 560078, India
    8. Department of Microbiology, Indian Academy Centre for Research and Postgraduate Studies, Bangalore 560040, India
  • Received:2011-07-02 Accepted:2011-08-29 Online:2011-11-20 Published:2011-11-15

Objective: The present study was undertaken to evaluate the effects of naringin on immobilization stress-induced biochemical-behavioral changes and mitochondrial dysfunction in mice.
Methods: Mice were randomized and grouped based on body weights. Respective drug treatments were given for 14 d, and on the 15th day all the animals were subjected to a 6-hour immobilization stress; then all the animals were subjected to various behavioral paradigms and were sacrificed. Various biochemical parameters and mitochondrial functions were analyzed using brain homogenate.
Results: The 6-hour acute immobilization stress significantly altered the behavioral (anxiety and memory) and biochemical parameters coupled with mitochondrial dysfunction in mice. Fourteen days pretreatment with naringin (50 and 100 mg/kg, per oral) significantly inhibited the behavioral and biochemical alterations and mitochondrial dysfunction caused by acute immobilization stress (P<0.05). Further, pretreatment with L-arginine (50 mg/kg, intraperitoneally), a nitric oxide precursor, reversed the protective effect of naringin (P<0.05). In addition, pretreatment with NG-nitro-L-arginine methyl ester (5 mg/kg, intraperitoneally) caused potentiation in the protective effect of naringin.
Conclusion: These results suggest the possible involvement of nitrergic pathway in the protective effect of naringin against immobilization stress-induced behavioral, biochemical and mitochondrial dysfunctions in mice.

Key words: naringin, restraint, physical, stress, nitric oxide, lipid peroxidation

Figure 1

Effects of naringin on IS-induced altered locomotor activity All the values are expressed as mean±SEM, n=10. *P<0.05, vs normal control; △P<0.05, vs IS control; ▲▲P<0.01, vs L-arginine (50 mg/kg) plus naringin (50 mg/kg); □P<0.05, vs L-arginine (50 mg/kg) plus naringin (100 mg/kg); ■P<0.05, vs L-arginine (50 mg/kg). IS: immobilization stress; Nar: naringin; SEM: standard error of mean; L-NAME: NG-nitro-L-arginine methyl ester."

Figure 2

Effects of naringin on IS-induced anxiety-like behavior observed by mirror chamber test (latency to enter into the mirror chamber) All the values are expressed as mean±SEM, n=10. *P<0.05, vs normal control; △P<0.05, vs IS control; ▲▲P<0.01, vs L-arginine (50 mg/kg) plus naringin (50 mg/kg); □P<0.05, vs L-arginine (50 mg/kg) plus naringin (100 mg/kg); ■P<0.05, vs L-arginine (50 mg/kg). IS: immobilization stress; Nar: naringin; SEM: standard error of mean; L-NAME: NG-nitro-L-arginine methyl ester."

Figure 3

Effects of naringin on IS-induced anxiety-like behavior observed by mirror chamber test (average time spent per entry in the mirror chamber) All the values are expressed as mean±SEM, n=10. *P<0.05, vs normal control; △P<0.05, vs IS control; ▲▲P<0.01, vs L-arginine (50 mg/kg) plus naringin (50 mg/kg); □P<0.05, vs L-arginine (50 mg/kg) plus naringin (100 mg/kg); ■P<0.05, vs L-arginine (50 mg/kg). IS: immobilization stress; Nar: naringin; SEM: standard error of mean; L-NAME: NG-nitro-L-arginine methyl ester."

Figure 4

Effects of naringin on IS-induced anxiety-like behavior observed by elevated plus-maze test (number of entries into the open arm) All the values are expressed as mean±SEM, n=10. *P<0.05, vs normal control; △P<0.05, vs IS control; ▲▲P<0.01, vs L-arginine (50 mg/kg) plus naringin (50 mg/kg); □P<0.05, vs L-arginine (50 mg/kg) plus naringin (100 mg/kg); ■P<0.05, vs L-arginine (50 mg/kg). IS: immobilization stress; Nar: naringin; SEM: standard error of mean; L-NAME: NG-nitro-L-arginine methyl ester."

Figure 5

Effects of naringin on IS-induced anxiety-like behavior observed by elevated plus-maze test (time spent in the open arm) All the values are expressed as mean±SEM, n=10. *P<0.05, vs normal control; △P<0.05, vs IS control; ▲▲P<0.01, vs L-arginine (50 mg/kg) plus naringin (50 mg/kg); □P<0.05, vs L-arginine (50 mg/kg) plus naringin (100 mg/kg); ■P<0.05, vs L-arginine (50 mg/kg). IS: immobilization stress; Nar: naringin; SEM: standard error of mean; L-NAME: NG-nitro-L-arginine methyl ester."




Reduced GSH
(μmol/mg protein)

(nmol/mg protein)

Catalase (μmol
H2O2/(min·mg protein))

(μmol/mg protein)

Normal control






IS control






Naringin (50 mg/kg)






Naringin (100 mg/kg)






L-arginine (50 mg/kg)






L-NAME (5 mg/kg)






L-arginine (50 mg/kg)
plus naringin (50 mg/kg)






L-arginine (50 mg/kg)
plus naringin (100 mg/kg)






L-NAME (5 mg/kg)
plus naringin (50 mg/kg)






L-NAME (5 mg/kg)
plus naringin (100 mg/kg)









Complex Ⅰ

Complex Ⅱ

MTT assay

Complex Ⅳ

Normal control






IS control






Naringin (50 mg/kg)






Naringin (100 mg/kg)






L-arginine (50 mg/kg)






L-NAME (5 mg/kg)






L-arginine (50 mg/kg)
plus naringin (50 mg/kg)






L-Arginine (50 mg/kg)
plus naringin (100 mg/kg)






L-NAME (5 mg/kg)
plus naringin (50 mg/kg)






L-NAME (5 mg/kg)
plus naringin (100 mg/kg)





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