Effects of long-term treatment with desipramine on microtubule proteins in rat cerebral cortex

doi:10.1016/S0014-2999(97)01140-0

European Journal of Pharmacology

Volume 333, Issues 2–3, 27 August 1997, Pages 279-287

https://doi.org/10.1016/S0014-2999(97)01140-0 Get rights and content

Abstract

The molecular mechanism of the action of antidepressants beyond the receptor level has not yet been elucidated. We have investigated the effects of long-term treatment with desipramine on the phosphorylation state of microtubule-associated protein 2 (MAP2) and microtubule assembly in the rat cerebral cortex. Phosphorylation of MAP2 was detected by immunoblotting after immunoprecipitation of MAP2 in the soluble fraction. The degree of phosphorylation of serine residues of MAP2 was significantly increased after chronic administration of desipramine without changes in the total concentration of MAP2. Microtubule assembly in crude brain extracts was monitored in terms of changes in turbidity measured at 350 nm using a spectrophotometer. Chronic but not acute treatment with desipramine inhibited microtubule assembly, assayed in the presence of a phosphatase inhibitor, calyculin A, whereas the inhibition was completely nullified in the absence of calyculin A. Desipramine had no direct effect on microtubule assembly in vitro. These results raise the possibility that the changes in the degree of phosphorylation of MAP2 and microtubule assembly represent intracellular modifications involved in functional changes elicited by long-term treatment with desipramine.

Introduction

The molecular mechanism of the action of antidepressants beyond the receptor level has not yet been elucidated. Previously, long-term treatment with desipramine, a tricyclic antidepressant drug, was reported to modify the endogenous phosphorylation of microtubule-associated protein 2 (MAP2) in both soluble and crude microtubule fractions of the rat cerebral cortex (Perez et al., 1989). MAP2 is known to be a substrate for several protein kinases such as type II cyclic AMP (cAMP)-dependent protein kinase (Vallee, 1980), Ca²⁺/calmodulin-dependent protein kinase II (Goldenring et al., 1983; Schulman, 1984) and Ca²⁺/phospholipid-dependent protein kinase (Akiyama et al., 1986). Several lines of evidence demonstrate that phosphorylation of MAP2 catalyzed by each of these protein kinases results in the inhibition of microtubule assembly in vitro (Jameson et al., 1980; Yamamoto et al., 1985; Hoshi et al., 1988). Thus, it is of interest to determine the effect of administration of antidepressants on microtubule assembly as a potential target for the action of antidepressants. In this study, we investigated the effects of long-term treatment with desipramine on the phosphorylation state of MAP2 as well as microtubule assembly in the rat cerebral cortex.

Section snippets

Antibodies

Monoclonal anti-MAP2 (2a-2b) (mouse IgG1) and anti-MAP1 (mouse IgG1) were obtained from Sigma Chemical (USA). Monoclonal anti-tau (mouse IgG1) was purchased from Chemicon International (USA). Monoclonal anti-α-tubulin (mouse IgG1) and anti-β-tubulin (mouse IgG1) were obtained from Oncogene Science (USA). Monoclonal anti-phosphoserine (mouse IgG1), anti-phosphothreonine (mouse IgG2b) and anti-phosphotyrosine (mouse IgG1) were obtained from Bio-Makor (Israel).

Animals and drug treatments

Adult male Wistar rats (150–200 g)

Effect of chronic desipramine treatment on MAPs phosphorylation

As shown in Fig. 1A (lanes 1 and 2), the anti-MAP2 monoclonal antibody stained a band of molecular weight 280 kDa of protein after immunoprecipitation of MAP2 from the cerebrocortical boiled supernatant fraction and electrophoresis of the immunoprecipitated MAP2. We observed no significant difference in the immunoreactivity of MAP2 between rats treated chronically with saline or desipramine (Fig. 1A and B, lanes 1 and 2). As shown in Fig. 1A and B (lanes 3 and 4), the immunoreactivity of the

Discussion

Although the therapeutic efficacy of antidepressants in the treatment of depression is well established, their precise mechanism of action has not yet been elucidated. A single administration of antidepressants induces an inhibition of monoamine reuptake within minutes to hours, whereas the clinical response to antidepressants usually requires 1 to 3 weeks to become evident (Heninger and Charney, 1987). The dissociation between the acute effect of antidepressants and the delayed onset of

Acknowledgements

We thank Ms. Yuriko Ogawa and Mr. Yuji Gamo for their assistance. This study was supported by a Research Grant for Nervous and Mental Disorders from the Ministry of Health and Welfare, Japan.

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Effects of long-term treatment with desipramine on microtubule proteins in rat cerebral cortex

Abstract

Introduction

Section snippets

Antibodies

Animals and drug treatments

Effect of chronic desipramine treatment on MAPs phosphorylation

Discussion

Acknowledgements

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

FEBS Lett.

J. Biol. Chem.

J. Biol. Chem.

FEBS Lett.

Neuropsychopharmacology

Neurosci. Lett.

J. Biol. Chem.

Eur. J. Pharmacol. Mol. Pharmacol. Sec.

Mol. Brain Res.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

Receptor sensitivity and the mechanism of action of antidepressant treatment

Arch. Gen. Psychiatry

Rapid dephosphorylation of microtubule-associated protein 2 in the rat brain hippocampus after pentylenetetrazole-induced seizures

Eur. J. Biochem.

An increase in phosphorylation of microtubule-associated protein 2 accompanies dendrite extension during the differentiation of cultured hippocampal neurones

Eur. J. Biochem.

Adaptations of receptor-coupled signal transduction pathways underlying stress- and drug-induced neural plasticity

J. Nerv. Ment. Dis.

Microtubule assembly in vitro

Eur. J. Biochem.

Phosphorylation of microtubule-associated protein 2 at distinct sites by calmodulin-dependent and cyclic-AMP-dependent kinases

J. Neurochem.

Microtubule destabilization and neurofilament phosphorylation precede dendritic sprouting after close axotomy of lamprey central neurones

Proc. Natl. Acad. Sci. U.S.A.

Participation of tubulin in the stimulatory regulation of adenylyl cyclase in rat cerebral cortex membranes

J. Neurochem.

Protein-kinase-C-catalyzed phosphorylation of the microtubule-binding domain of microtubule-associated protein 2 inhibits its ability to induce tubulin polymerization

Eur. J. Biochem.

A comparative study of the peroxidase–antiperoxidase method and an avidin–biotin complex method for studying polypeptide hormones with radioimmunoassay antibodies

Am. J. Clin. Pathol.