Bee venom and its component apamin as neuroprotective agents in a Parkinson disease mouse model

Abstract

Bee venom has recently been suggested to possess beneficial effects in the treatment of Parkinson disease (PD). For instance, it has been observed that bilateral acupoint stimulation of lower hind limbs with bee venom was protective in the acute 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of PD. In particular, a specific component of bee venom, apamin, has previously been shown to have protective effects on dopaminergic neurons in vitro. However, no information regarding a potential protective action of apamin in animal models of PD is available to date. The specific goals of the present study were to (i) establish that the protective effect of bee venom for dopaminergic neurons is not restricted to acupoint stimulation, but can also be observed using a more conventional mode of administration and to (ii) demonstrate that apamin can mimic the protective effects of a bee venom treatment on dopaminergic neurons. Using the chronic mouse model of MPTP/probenecid, we show that bee venom provides sustained protection in an animal model that mimics the chronic degenerative process of PD. Apamin, however, reproduced these protective effects only partially, suggesting that other components of bee venom enhance the protective action of the peptide.

Figure 1. Time schedule for bee venom and apamin injections in MPTP/probenecid-intoxicated mice.

Time points at which MPTP/probenecid (red arrows) and bee venom or apamin (green arrows) injections were carried out. Note that bee venom and apamin were given every 3.5 days, starting 48 h day after the first MPTP/probenecid injection. Blue and grey arrows depict time points at which mice were taken for the open field or rotarod testing, respectively. Habituation to the open field test and training to the rotarod test were carried out 3 and 2 days before beginning the first MPTP/probenecid injection, respectively. Baseline values for the open field and rotarod tests were acquired one day before, and on the same day as the first injection with MPTP/probenecid, respectively. For the rotarod testing, post-treatment values were obtained one day after the ninth injection with bee venom or apamin and for the open field testing 120 min after the last injection with of one of these two treatments. After the open field testing, all mice were killed for brain processing and analysis.

Figure 2. Effect of bee venom (Alyo) and apamin (Apam) treatments against MPTP intoxication.

(A) Number of SNpc TH+ cells of the right hemisphere in MPTP/probenecid-treated mice receiving or not treatments with bee venom or apamin. Legends corresponding to various treatment paradigms are given above. (B) Striatal DA levels and (C) HVA/DA ratios in mice receiving the same treatment regimens. Data are expressed as percentage of saline-injected mice. Actual values for controls were as follows: TH⁺ cell numbers 5112.5±203.3, striatal DA levels, 8.82 ng/mg wet tissue ±0.42; HVA/DA ratios, 0.18±0.01. Data represent mean values ± s.e.m of 5–8 animals. *p<0.05 compared to control values, #p<0.05 for pairwise comparison.

Figure 3. Behavioural effects of bee venom (Alyo) and apamin (Apam) in mice exposed or not to repeated MPTP/probenecid injections.

(A) Track length reached in the open field after 25 min (100% = 6998 cm±226) and (B) rotarod performance (100% = 240 seconds) as a function of the treatments mentioned above. Baseline values (prae) were obtained at 10 rotations per minute (rpm). Post-treatment values were obtained both at 10 and 20 rpm after the ninth bee venom/apamin/vehicle injection (Fig. 1). Data are expressed as mean ± s.e.m. *p<0.05 compared to control values, #p<0.05 for pairwise comparison.

Figure 4. Effects of bee venom (Alyo) and apamin (Apam) on striatal cytokine levels in mice exposed to repeated MPTP/probenecid vs. saline injections.

(A) IL-1β, (B) IL-6 and (C) TNF-α striatal levels. Data are expressed as mean ± s.e.m. *p<0.05 compared to control values, #p<0.05 for pairwise comparison.

Figure 5. Effects of bee venom (Alyo) and apamin (Apam) on striatal Cx I activity in mice exposed to repeated MPTP/probenecid vs. saline injections.

The ratio of Cx I/CS activity is given in the graph as percentage of saline/saline animals with n = 4 per group. Analysis was performed in four independent experiments whereas in each experiment the obtained value of the saline/saline group (expressed as nmol/ml*min) was set to 100%. Data are expressed as mean ± s.e.m. *p<0.05 compared to control values, #p<0.05 for pairwise comparison.