Abstract
Protein phosphorylation was examined in the identified Aplysia neuron R15, in vivo, after the intracellular injection of [γ-32P]ATP. Two-dimensional gel electrophoretic analysis indicates that at least 70 proteins are phosphorylated within R15 during a 50-min labeling period. Application of serotonin (5HT) results in an increase in K+ conductance in R15 and a concomitant change in the phosphorylation pattern: there are increases or decreases in the phosphorylation of some proteins, and at least five phosphoproteins appear that are not observed in control cells. Dopamine causes a decrease in voltage-dependent inward conductance in R15 and also alters the phosphorylation pattern: several of the phosphorylation changes are similar to those produced by 5HT, while others are unique to dopamine. Stimulation of the branchial nerve leading to the abdominal ganglion results in a long-lasting synaptic hyperpolarization of R15. The conductance changes underlying this response include an increase in K+ conductance (identical to that produced by 5HT) together with a decrease in voltage-dependent inward conductance (identical to that produced by dopamine). The phosphorylation changes induced in R15 by branchial nerve stimulation resemble a combination of the changes induced by 5HT and dopamine. The results demonstrate that synaptic stimulation can modulate the phosphorylation of specific proteins in a single identified postsynaptic neuron and are consistent with the hypothesis that protein phosphorylation can regulate the regulate the activity of neuronal ion channels.
Keywords: neurotransmitters, ion conductance, synaptic transmission, cAMP, single-cell phosphorylation
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- Adams W. B., Levitan I. B. Intracellular injection of protein kinase inhibitor blocks the serotonin-induced increase in K+ conductance in Aplysia neuron R15. Proc Natl Acad Sci U S A. 1982 Jun;79(12):3877–3880. doi: 10.1073/pnas.79.12.3877. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Adams W. B., Parnas I., Levitan I. B. Mechanism of long-lasting synaptic inhibition in Aplysia neuron R15. J Neurophysiol. 1980 Dec;44(6):1148–1160. doi: 10.1152/jn.1980.44.6.1148. [DOI] [PubMed] [Google Scholar]
- Ascher P. Inhibitory and excitatory effects of dopamine on Aplysia neurones. J Physiol. 1972 Aug;225(1):173–209. doi: 10.1113/jphysiol.1972.sp009933. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bandle E. F., Levitan I. B. Cyclic AMP-stimulated phosphorylation of a high molecular weight endogenous protein substrate in sub-cellular fractions of molluscan nervous system. Brain Res. 1977 Apr 15;125(2):325–331. doi: 10.1016/0006-8993(77)90625-4. [DOI] [PubMed] [Google Scholar]
- Benson J. A., Levitan I. B. Serotonin increases an anomalously rectifying K+ current in the Aplysia neuron R15. Proc Natl Acad Sci U S A. 1983 Jun;80(11):3522–3525. doi: 10.1073/pnas.80.11.3522. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bernier L., Castellucci V. F., Kandel E. R., Schwartz J. H. Facilitatory transmitter causes a selective and prolonged increase in adenosine 3':5'-monophosphate in sensory neurons mediating the gill and siphon withdrawal reflex in Aplysia. J Neurosci. 1982 Dec;2(12):1682–1691. doi: 10.1523/JNEUROSCI.02-12-01682.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Castellucci V. F., Kandel E. R., Schwartz J. H., Wilson F. D., Nairn A. C., Greengard P. Intracellular injection of t he catalytic subunit of cyclic AMP-dependent protein kinase simulates facilitation of transmitter release underlying behavioral sensitization in Aplysia. Proc Natl Acad Sci U S A. 1980 Dec;77(12):7492–7496. doi: 10.1073/pnas.77.12.7492. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Castellucci V. F., Nairn A., Greengard P., Schwartz J. H., Kandel E. R. Inhibitor of adenosine 3':5'-monophosphate-dependent protein kinase blocks presynaptic facilitation in Aplysia. J Neurosci. 1982 Dec;2(12):1673–1681. doi: 10.1523/JNEUROSCI.02-12-01673.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Deterre P., Paupardin-Tritsch D., Bockaert J., Gerschenfeld H. M. Role of cyclic AMP in a serotonin-evoked slow inward current in snail neurones. Nature. 1981 Apr 30;290(5809):783–785. doi: 10.1038/290783a0. [DOI] [PubMed] [Google Scholar]
- Drummond A. H., Benson J. A., Levitan I. B. Serotonin-induced hyperpolarization of an indentified Aplysia neuron is mediated by cyclic AMP. Proc Natl Acad Sci U S A. 1980 Aug;77(8):5013–5017. doi: 10.1073/pnas.77.8.5013. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Greengard P. Cyclic nucleotides, phosphorylated proteins, and the nervous system. Fed Proc. 1979 Jul;38(8):2208–2217. [PubMed] [Google Scholar]
- Jennings K. R., Kaczmarek L. K., Hewick R. M., Dreyer W. J., Strumwasser F. Protein phosphorylation during afterdischarge in peptidergic neurons of Aplysia. J Neurosci. 1982 Feb;2(2):158–168. doi: 10.1523/JNEUROSCI.02-02-00158.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kaczmarek L. K., Jennings K. R., Strumwasser F., Nairn A. C., Walter U., Wilson F. D., Greengard P. Microinjection of catalytic subunit of cyclic AMP-dependent protein kinase enhances calcium action potentials of bag cell neurons in cell culture. Proc Natl Acad Sci U S A. 1980 Dec;77(12):7487–7491. doi: 10.1073/pnas.77.12.7487. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kaczmarek L. K., Jennings K., Strumwasser F. Neurotransmitter modulation, phosphodiesterase inhibitor effects, and cyclic AMP correlates of afterdischarge in peptidergic neurites. Proc Natl Acad Sci U S A. 1978 Oct;75(10):5200–5204. doi: 10.1073/pnas.75.10.5200. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Klein M., Kandel E. R. Mechanism of calcium current modulation underlying presynaptic facilitation and behavioral sensitization in Aplysia. Proc Natl Acad Sci U S A. 1980 Nov;77(11):6912–6916. doi: 10.1073/pnas.77.11.6912. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kuo J. F., Greengard P. Cyclic nucleotide-dependent protein kinases. IV. Widespread occurrence of adenosine 3',5'-monophosphate-dependent protein kinase in various tissues and phyla of the animal kingdom. Proc Natl Acad Sci U S A. 1969 Dec;64(4):1349–1355. doi: 10.1073/pnas.64.4.1349. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Lemos J. R., Levitan I. B. Intracellular injection of guanyl nucleotides alters the serotonin-induced increase in potassium conductance in Aplysia neuron R15. J Gen Physiol. 1984 Feb;83(2):269–285. doi: 10.1085/jgp.83.2.269. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Lemos J. R., Novak-Hofer I., Levitan I. B. Serotonin alters the phosphorylation of specific proteins inside a single living nerve cell. Nature. 1982 Jul 1;298(5869):64–65. doi: 10.1038/298064a0. [DOI] [PubMed] [Google Scholar]
- Levitan I. B., Barondes S. H. Octopamine- and serotonin-stimulated phosphorylation of specific protein in the abdominal ganglion of Aplysia californica. Proc Natl Acad Sci U S A. 1974 Apr;71(4):1145–1148. doi: 10.1073/pnas.71.4.1145. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Levitan I. B., Madsen C. J., Barondes S. H. Cyclic AMP and amine effects on phosphorylation of specific protein in abdominal ganglion of Aplysia californica; localization and kinetic analysis. J Neurobiol. 1974;5(6):511–525. doi: 10.1002/neu.480050604. [DOI] [PubMed] [Google Scholar]
- Novak-Hofer I., Levitan I. B. Ca++/calmodulin-regulated protein phosphorylation in the Aplysia nervous system. J Neurosci. 1983 Mar;3(3):473–481. doi: 10.1523/JNEUROSCI.03-03-00473.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
- O'Farrell P. H. High resolution two-dimensional electrophoresis of proteins. J Biol Chem. 1975 May 25;250(10):4007–4021. [PMC free article] [PubMed] [Google Scholar]
- Osterrieder W., Brum G., Hescheler J., Trautwein W., Flockerzi V., Hofmann F. Injection of subunits of cyclic AMP-dependent protein kinase into cardiac myocytes modulates Ca2+ current. Nature. 1982 Aug 5;298(5874):576–578. doi: 10.1038/298576a0. [DOI] [PubMed] [Google Scholar]
- Parnas I., Armstrong D., Strumwasser F. Prolonged excitatory and inhibitory synaptic modulation of a bursting pacemaker neuron. J Neurophysiol. 1974 Jul;37(4):594–608. doi: 10.1152/jn.1974.37.4.594. [DOI] [PubMed] [Google Scholar]
- Parnas I., Strumwasser F. Mechanisms of long-lasting inhibition of a bursting pacemaker neuron. J Neurophysiol. 1974 Jul;37(4):609–620. doi: 10.1152/jn.1974.37.4.609. [DOI] [PubMed] [Google Scholar]
- Pellmar T. C. Ionic mechanism of a voltage-dependent current elicited by cyclic AMP. Cell Mol Neurobiol. 1981 Mar;1(1):87–97. doi: 10.1007/BF00736041. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Stein C., Weinreich D. Determination of glutathione and ATP in ganglia and individual neurons of Aplysia californica. J Neurochem. 1984 Apr;42(4):1170–1174. doi: 10.1111/j.1471-4159.1984.tb12726.x. [DOI] [PubMed] [Google Scholar]
- Tsien R. W. Adrenaline-like effects of intracellular iontophoresis of cyclic AMP in cardiac Purkinje fibres. Nat New Biol. 1973 Sep 26;245(143):120–122. doi: 10.1038/newbio245120a0. [DOI] [PubMed] [Google Scholar]
- Wilson W. A., Wachtel H. Prolonged inhibition in burst firing neurons: synaptic inactivation of the slow regenerative inward current. Science. 1978 Nov 17;202(4369):772–775. doi: 10.1126/science.715442. [DOI] [PubMed] [Google Scholar]
- de Peyer J. E., Cachelin A. B., Levitan I. B., Reuter H. Ca2+ -activated K+ conductance in internally perfused snail neurons is enhanced by protein phosphorylation. Proc Natl Acad Sci U S A. 1982 Jul;79(13):4207–4211. doi: 10.1073/pnas.79.13.4207. [DOI] [PMC free article] [PubMed] [Google Scholar]






