Reversible Downregulation of Protein Kinase A during Olfactory Learning Using Antisense Technique Impairs Long-Term Memory Formation in the Honeybee, Apis mellifera
Reversible Downregulation of Protein Kinase A during Olfactory Learning Using Antisense Technique Impairs Long-Term Memory Formation in the Honeybee, Apis mellifera
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DOI：
10.1523/jneurosci.19-22-10125.1999
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发表时间：
1999-11
期刊：
影响因子：
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通讯作者：
A. Fiala;U. Müller;R. Menzel
中科院分区：
文献类型：
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作者：
A. Fiala;U. Müller;R. Menzel
在这项研究中，我们研究了cAMP依赖性蛋白激酶（PKA）在蜜蜂，意大利蜜蜂的联想嗅觉学习中的作用。在蜜蜂中，对分子进行特异性干扰以阐明它们在某种行为中的作用是困难的，因为遗传方法，如突变体或转基因动物，目前还不可行。作为一种新的方法在昆虫体内，我们报告使用短的反义寡核苷酸。我们表明，硫代磷酸修饰的寡脱氧核苷酸互补的PKA的催化亚基的mRNA直接注射到蜜蜂的大脑引起可逆的和特定的下调的催化亚基和PKA活性的量的10- 15%。PKA的调节亚基以及PKC的量不受影响。轻微的“击倒”PKA活性在训练过程中，一个经典的嗅觉条件反射的长鼻延伸，既不影响收购，也不影响记忆保留3或6小时后的培训。然而，在训练后24小时，它会导致长期记忆保留的损害。训练后3小时PKA的下调对记忆形成没有可检测的影响。我们的结论是PKA有助于诱导的长期记忆24小时后，在学习过程中激活培训。其次，我们证明了反义技术在蜜蜂体内是可行的，并为昆虫学习记忆形成的分子基础的研究提供了一个新的和强有力的工具。
In this study, we examined the role of cAMP-dependent protein kinase (PKA) in associative olfactory learning of the honeybee,Apis mellifera. In the bee, specific interference with molecules to clarify their role in a certain behavior is difficult, because genetic approaches, such as mutants or transgenic animals, are not feasible at the moment. As a new approach in insects in vivo, we report the use of short antisense oligonucleotides. We show that phosphorothioate-modified oligodeoxynucleotides complementary to the mRNA of a catalytic subunit of PKA directly injected into the bee brain cause a reversible and specific downregulation of both the amount of the catalytic subunit and of PKA activity by 10–15%. The amounts of the regulatory subunit of PKA, as well as PKC, are not affected. The slight “knockdown” of PKA activity during the training procedure, a classical olfactory conditioning of the proboscis extension reflex, neither affects acquisition nor memory retention 3 or 6 hr after training. However, it causes an impairment of long-term memory retention 24 hr after training. Downregulation of PKA 3 hr after training has no detectable effect on memory formation. We conclude that PKA contributes to the induction of a long-term memory 24 hr after training when activated during learning. Second, we demonstrate that the antisense technique is feasible in honeybees in vivo and provides a new and powerful tool for the study of the molecular basis of learning and memory formation in insects.