Zinc Deficiency Of CD-1 Mice During Lactation Period Causes Neuronal Apoptosis In The Hippocampus Of Their Suckling Offsprings

PrefaceZinc is an important trace element and is required for several cellular processes such as gene expression,DNA synthesis,enzymatic catalysis,tissue repair, neurotransmission and memory formation.The majority of zinc in the brain (approximately 85%) is tightly bound to proteins,such as zinc finger motifs, metallothioneins,while the rest(approximately 15%) is sequestered in presynaptic vesicles in the terminal boutons of zinc-containing neurons.It has been proposed that zinc ions released into the synaptic clefts act as a neuromodulator to modulate postsynaptic receptors,including NMDA,AMPA/KA and GABA receptors.Many studies indicate that zinc-releasing synapses might have a special role in the synaptic plasticity that underlies learning and memory.Zinc ions have also been suggested to inhibit apoptosis and promote mitosis. Accumulating evidence has demonstrated that zinc deficiency causes neuronal apoptosis.The mechanism,however,is far from fully understood.The vesicular zinc is able to regulate the activity of TrkB via BDNF or Src.Meanwhile ERK,which is a downstream protein of TrkB,can modulate many apoptosis-regulatory proteins like Bax,Bcl-2 and Caspase-3.Therefore,in the present study,we focused on the effects of zinc deficiency on TrkB-related pathways,and aimed to explore the mechanisms underlying neuronal apoptosis in hippocampus caused by dietary zinc deficiency.Materials and MethodsPregnant CD-1 mice were randomly assigned to one of the two dietary groups (n=6) in each group:zinc-adequate group or control group and zinc-deficient group.All mice were housed in stainless steel cages.From the 1^st day after giving birth,mothers in experimental groups were fed with zinc-deficient diet(zinc content is 0.85ppm) and deionized water.Meanwhile animals in control groups were fed with normal diet(zinc content is 30mg/kg) and deionized water.Mice were sacrificed at three time points to get hippocampus including P7,P14 and P21.Autometallography(AMG),Nissl staining, TUNEL staining and Western blot were used to analyze the changes of vesicular zinc ions,neuronal apoptosis and the levels of related proteins in hippocampus of sucking mice.ResultsAMG results showed that the content of free zinc ions in the hippocampus of zinc-deficient animals was obviously lower than that of normal animals during sucking period.Nissl and TUNEL staining demonstrated that more neuronal loss and apoptosis existed in the hippocampus of zinc-deficient offspring.Furthermore,Western blot revealed the higher expressions of pro-BDNF(28kDa) and BDNF(14kDa) in the hippocampus of the zinc-deficient mice at all the three time points(P7,P14,P21).At the same time,the levels of Src made no difference between control and experimental groups but that of p-Src(Tyr527) increased due to dietary zinc deficiency.The expressions of p-TrkB decreased in the hippocampus of zinc-deficient sucking mice while the total content of TrkB did not change.ERK contains two types including ERK1 and ERK2(P44 and P42 respectively),which are the key factors transferring signals from surface receptors to nuclei.The total content of ERK did not affected by zinc deficiency,but the levels of p-ERK decreased.As to apoptosis-regulatory proteins, we found that Bax/Bcl-2 and Caspase-3 were significantly increased in zinc-deficient mice.ConclusionZinc deficiency occurred to lactating mice can lead to a decrease in free zinc ions and an increase in neuronal apoptosis in the hippocampus of their offspring during sucking period.Based on the results above,we propose a possible mechanism that can explain the phenomenon.That is,zinc deficiency inhibits Src/TrkB/ERK pathway and then promotes the expressions of apoptosis-inducing proteins including Bax and Caspase-3 and reduce the expression of anti-apoptotic protein like Bcl-2.Moreover, higher expression of BDNF is considered as a protective response,which cannot fully compensate the injury caused by zinc deficiency.