Inositol has behavioral effects with adaptation after chronic administration

Summary Inositol is a simple dietary polyol that serves as a precursor in important second messenger systems. Inositol in pharmacological doses has been reported recently to be therapeutic in depression, panic disorder and obsessive compulsive disorder. We hereby report effects of inositol in the elevated plus maze model of anxiety. These results should allow development of new inositol analogs that could expand psychoactive drug development possibilities via second messenger manipulation.     

Lithium-pilocarpine seizures as a model for lithium action in mania

Lithium (Li) pre-treatment of rats or mice given low dose pilocarpine induces a unique limbic seizure syndrome. This syndrome is stereospecifically reversed by myo-inositol, which suggests that it is a behavioral model for Li depletion of brain inositol. However, this syndrome has little face validity because seizures are not a component of bipolar disorder. Moreover, other animal species that maintain higher brain inositol levels than mice or rats do not show Li-pilocarpine seizures and a study in humans suggests that humans do not show this syndrome as well. It could be suggested that Li-pilocarpine seizures are an in vivo bioassay for inositol depletion. Recent studies of knockout mice lacking inositol monophosphatase-1 or the sodium myo-inositol transporter-1 found that both these knockout mice given pilocarpine develop limbic seizures as if they had been pre-treated with Li. These mice in addition to such pilocarpine sensitivity have other behaviors such as decreased immobility in the Porsolt forced swim test that suggests that their inositol depletion has Li-like effects. Thus, the Li-pilocarpine seizure model may, despite its lack of face validity, be a biochemical marker for a model of mania treatment in animals.     

Lithium alters regional rat brain myo-inositol at 2 and 4 weeks: an ex-vivo magnetic resonance spectroscopy study at 18.8 T

Lithium has been the mainstay of treatment for bipolar disorder. Early studies suggest that lithium acts via inositol depletion. This study assesses the effect of 1, 2 and 4 weeks of lithium treatment on myo-inositol concentrations across several brain regions. Thirty-six Sprague-Dawley rats were treated for 2 weeks with an intraperitoneal injection of either 1 mmol/kg/day, twice daily lithium chloride (n=18) or placebo (2 ml/kg of saline) (n=18). The rats were separated into three groups: 1 week, 2 weeks and 4 weeks. Brains were dissected into prefrontal, temporal and occipital cortical areas, as well as hippocampus, and analyzed at 18.8 T. Myo-inositol was quantified using the Chenomx Profiler software. Lithium did not alter myo-inositol concentrations at 1 week. A significant reduction exists in myo-inositol concentrations in lithium-treated rats at 2 and 4 weeks, across all four brain regions. Studies suggest brain region-specific alterations in myo-inositol concentrations among bipolar patients. Our findings suggest that lithium-induced reduction of myo-inositol is more global.     

The anxiolytic effect of chronic inositol depends on the baseline level of anxiety

Inositol, a precursor for membrane phosphoinositides involved in signal transduction, has been found to be clinically effective in a number of psychiatric disorders and to reverse behavioural effects of lithium. To gain insight into the mechanism of action of inositol, it is critical to establish its efficacy in animal models. Following the initial report by Cohen et al. (1997b) that inositol was anxiolytic in the elevated plus maze model of anxiety, the effect of chronic intraperitoneal and chronic dietary inositol administration in rats was tested in four experiments. There was a significant increase in closed arm and total arm entries following chronic injection of inositol, but no effect of inositol when it was given chronically in rat chow. Because the first 2 experiments suggested that the mode of drug administration affected the control levels of anxiety (open arm entries and time in open arms) in control groups, the effect of chronic dietary inositol was tested in rats that were exposed to a mild and a more severe form of stress. Chronic saline injections elevated anxiety in the plus maze, which was only marginally affected by chronic dietary inositol. Following 3 weeks administration of 5% dietary inositol rats were pre-exposed to a cat. There was a clear increase in number of entries into open arms, suggesting an anxiolytic effect of inositol.     

Myo-inositol has no beneficial effect on premenstrual dysphoric disorder.

Inositol, a simple isomer of glucose, which serves as a precursor in the phosphatidyl-inositol (PI) second messenger cycle, was shown to be effective in double-blind, placebo-controlled studies of depression, panic and obsessive compulsive disorders as well as in bulimia. The following study was designed to investigate whether inositol has beneficial effects in another disorder shown to be responsive to SSRIs: premenstrual dysphoric disorder (PMDD). Eleven female patients with PMDD diagnosed according to DSM-IV participated in a cross-over, double-blind, placebo-controlled trial. The active drug was myo-inositol, 12 g daily, whereas placebo was d-glucose administered at the same dose. Each drug was given during the luteal phase only (14 days prior to menses). For each patient treatment alternated between these two drugs for six menstrual cycles. No beneficial effect was demonstrated for inositol over placebo.     

Temporal dissociation between lithium-induced changes in frontal lobe myo-inositol and clinical response in manic-depressive illness

OBJECTIVE: The most widely accepted hypothesis regarding the mechanism underlying lithium's therapeutic efficacy in manic-depressive illness (bipolar affective disorder) is the inositol depletion hypothesis, which posits that lithium produces a lowering of myo-inositol in critical areas of the brain and the effect is therapeutic. Lithium's effects on in vivo brain myo-inositol levels were investigated longitudinally in 12 adult depressed patients with manic-depressive illness. METHOD: Medication washout (minimum 2 weeks) and lithium administration were conducted in a blinded manner. Regional brain myo-inositol levels were measured by means of quantitative proton magnetic resonance spectroscopy at three time points: at baseline and after acute (5-7 days) and chronic (3-4 weeks) lithium administration. RESULTS: Significant decreases (approximately 30%) in myoinositol levels were observed in the right frontal lobe after short-term administration, and these decreases persisted with chronic treatment. The severity of depression measured by the Hamilton Depression Rating Scale also decreased significantly over the study. CONCLUSIONS: This study demonstrates that lithium administration does reduce myo-inositol levels in the right frontal lobe of patients with manic-depressive illness. However, the acute myo-inositol reduction occurs at a time when the patient's clinical state is clearly unchanged. Thus, the short-term reduction of myo-inositol per se is not associated with therapeutic response and does not support the inositol depletion hypothesis as originally posited. The hypothesis that a short-term lowering of myo inositol results in a cascade of secondary signaling and gene expression changes in the CNS that are ultimately associated with lithium's therapeutic efficacy is under investigation.     

Inositol treatment of autism

Summary Recent studies suggest that serotonin reuptake inhibitors are helpful in at least some symptoms of autism. Inositol is a precursor of the second messenger for some serotonin receptors, and has been reported effective in depression, panic disorder and obsessive-compulsive disorder. However a controlled double-blind crossover trial of inositol 200 mg/kg per day showed no benefit in 9 children with autism. Since biochemical studies suggest that inositol may augment serotonin effects, future studies could evaluate inositol in children already receiving serotonin reuptake inhibitors.     

Inositol 1,4,5-trisphosphate receptor binding: autoradiographic localization in rat brain

Inositol 1,4,5-trisphosphate is a second messenger generated by stimulation of the phosphoinositide cycle, thought to release calcium from intracellular stores. We have mapped the distribution of 3H-inositol 1,4,5-trisphosphate receptor binding sites in rat brain by autoradiographic techniques. The cerebellum contains the highest level of inositol 1,4,5-trisphosphate binding sites in brain, which appear to be selectively localized to Purkinje cells. Moderate levels of binding sites are present in the hippocampus, cerebral cortex, caudate, and substantia nigra. Lesion studies indicate that binding in the hippocampus is restricted to intrinsic neuronal elements and in the nigra is found on terminals of the striatonigral projection. Overall, the autoradiographic distribution of inositol 1,4,5-trisphosphate receptors resembles the distribution of phorbol ester binding sites associated with protein kinase C. However, the inositol 1,4,5-trisphosphate receptor has a more restricted distribution since it is not detectable in the spinal cord or olfactory bulb, regions with substantial levels of protein kinase C.     

Inositol deficiency diet and lithium effects

OBJECTIVES: A major hypothesis explaining the therapeutic effect of lithium (Li) in mania is depletion of inositol via inhibition of inositol monophosphatase. However, inositol is also present in the diet. Restriction of dietary inositol could theoretically enhance the effects of Li. METHODS: We used dietary inositol restriction in animal studies and also devised a palatable diet for humans that is 90% free of inositol. RESULTS: Dietary inositol restriction significantly augmented the inositol-reducing effect of Li in rat frontal cortex. Li reduced inositol levels by 4.7%, inositol-deficient diet by 5.1%, and Li plus inositol-deficient diet by 10.8%. However, feeding with the inositol-deficient diet did not enhance the behavioral effect of Li in the Li-pilocarpine seizure model. Fifteen patients participated in an open clinical study of the inositol-deficient diet: six rapid cycling bipolar patients responding inadequately to Li or valproate in different phases of illness; two Li-treated bipolar outpatients with residual symptomatology, and seven inpatient Li-treated bipolar patients in non-responding acute mania. The diet had a major effect in reducing the severity of affective disorder in 10 of the patients within the first 7-14 days of treatment. CONCLUSION: These results suggest that dietary inositol restriction may be useful in some bipolar patients, but controlled replication is necessary.     

Evidence that a proconvulsant action of lithium is mediated by inhibition of myo-inositol phosphatase in mouse brain.

Lithium inhibits myo-inositol mono- and polyphosphatase activity in brain at concentrations similar to those optimal for the treatment of manic depressive psychosis. A consequence of this inhibition is the possibility that the availability of myo-inositol for the regeneration of polyphosphoinositides involved in cellular signalling mechanisms may be reduced. While there are no good models of manic depressive disorders in rodents, lithium is known to alter their behavioural responsiveness to a number of neurotransmitter receptor agonists, but the role of the phosphatidylinositol second messenger system in these effects is unknown. Consistent with the myo-inositol depletion hypothesis, when injected directly into the CNS, myo-inositol, but not its biologically inactive epimer, scyllo-inositol or D-mannitol, has been found to reverse a proconvulsant action of lithium in mice given the muscarinic receptor agonist, pilocarpine.     

Unlike lithium, anticonvulsants and antidepressants do not alter rat brain myo-inositol

Lithium is the first-line in bipolar disorder treatment. Lithium's clinical efficacy might be due to its inhibition of myo-inositol turnover in the phosphatidylinositol second messenger system. This study aimed to determine whether this action can extend to antidepressants and anticonvulsants also used to treat bipolar symptoms. Male rats were treated for 2 weeks with an intraperitoneal injection of phenelzine, fluoxetine, desipramine, carbamazepine, lamotrigine, sodium valproate or vehicle. Brains were dissected and myo-inositol concentrations were analyzed using high-field nuclear magnetic resonance spectroscopy at 18.8 T and quantified using Chenomx Profiler software. Brain regions assessed included the prefrontal, temporal and occipital cortical areas as well as the hippocampus. The main finding is that contrary to lithium, the anticonvulsants and antidepressants do not alter brain myo-inositol concentration. This suggests that these agents might work via a mechanism that is not centered on changes in myo-inositol concentration.     

Acute myo-inositol enhances swimming activity in goldfish

Summary. Inositol in concentrations of 1–4 Mm (but not the control condition: mannitol-glucose) administered in aquarium water showed enhancement of swimming activity of goldfish after acute treatment (5 hours). These data support similar findings in rats. These data suggest that inositol may enhance motor activity which may be of relevance in neuropsychiatric disorders. Future studies of the effects of lithium on this activity, and its potential reversal by inositol may shed light on the possible involvement of the phosphatidyl-inositol second messenger system in this behavior. Received December 15, 1998; accepted January 20, 1999     

Bipolar disorder and myo-inositol: a review of the magnetic resonance spectroscopy findings

OBJECTIVES: Myo-inositol is an important component of the phosphatidylinositol second messenger system (PI-cycle). Alterations in PI-cycle activity have been suggested to be involved in the pathophysiology and/or treatment of bipolar disorder. More specifically, lithium has been suggested to act primarily by lowering myo-inositol concentrations, the so-called inositol-depletion hypothesis. myo-Inositol concentrations can be measured in vivo with magnetic resonance spectroscopy (MRS). METHODS: The current review primarily examines animal and human MRS studies that evaluated the role of myo-inositol in bipolar illness and treatment. RESULTS: Studies have been carried out in patients who are manic, depressed, and euthymic, both on and off treatment. However, there are several limitations of these studies. CONCLUSIONS: The preclinical and clinical MRS findings were generally supportive of the involvement of myo-inositol in bipolar disorder and its treatment. Overall, in bipolar patients who are manic or depressed there are abnormalities in brain myo-inositol concentrations, with changes in frontal and temporal lobes, as well as the cingulate gyrus and basal ganglia. These abnormalities are not seen in either euthymic patients or healthy controls, possibly due to a normalizing effect of treatment with either lithium or sodium valproate. There is also increasing evidence that sodium valproate may also act upon the PI-cycle. Nonetheless, it remains uncertain if these changes in myo-inositol concentration are primary or secondary. Findings regarding the specific inositol-depletion hypothesis are also generally supportive in acutely ill patients, although it is not yet possible to definitively confirm or refute this hypothesis based on the current MRS evidence.     

Valproate decreases inositol biosynthesis.

BACKGROUND: Lithium and valproate (VPA) are used for treating bipolar disorder. The mechanism of mood stabilization has not been elucidated, but the role of inositol has gained substantial support. Lithium inhibition of inositol monophosphatase, an enzyme required for inositol recycling and de novo synthesis, suggested the hypothesis that lithium depletes brain inositol and attenuates phosphoinositide signaling. Valproate also depletes inositol in yeast, Dictyostelium, and rat neurons. This raised the possibility that the effect is the result of myo-inositol-1-phosphate (MIP) synthase inhibition. METHODS: Inositol was measured by gas chromatography. Human prefrontal cortex MIP synthase activity was assayed in crude homogenate. INO1 was assessed by Northern blotting. Growth cones morphology was evaluated in cultured rat neurons. RESULTS: We found a 20% in vivo reduction of inositol in mouse frontal cortex after acute VPA administration. As hypothesized, inositol reduction resulted from decreased MIP synthase activity: .21-.28 mmol/LVPA reduced the activity by 50%. Among psychotropic drugs, the effect is specific to VPA. Accordingly, only VPA upregulates the yeast INO1 gene coding for MIP synthase. The VPA derivative N-methyl-2,2,3,3,-tetramethyl-cyclopropane carboxamide reduces MIP synthase activity and has an affect similar to that of VPA on rat neurons, whereas another VPA derivative, valpromide, poorly affects the activity and has no affect on neurons. CONCLUSIONS: The rate-limiting step of inositol biosynthesis, catalyzed by MIP synthase, is inhibited by VPA; inositol depletion is a first event shown to be common to lithium and VPA.     

Biological predictors of lithium response in bipolar disorder

In order to prescribe lithium appropriately to patients with bipolar disorder, predictors of lithium response are helpful. The present paper reviews the biological predictors of lithium response. As a positive predictor of lithium response, the following have been reported: strong loudness dependence of the auditory-evoked N1/P2-response; higher brain lithium concentration; lower inositol monophosphatase (IMPase) mRNA expression; higher serotonin-induced calcium mobilization; increased N-acetyl-aspartate peak and decreased myo-inositol peak; white matter hyperintensity; decreased intracellular pH; higher frequency of phospholipase C gamma-1 (PLCG1)-5 repeat and PLCG1-8 repeat; and C973A polymorphism in the inositol polyphosphate 1-phosphatase gene. In contrast the following have been reported as a predictor of negative lithium response: epileptiform abnormality of electroencephalography; human leukocyte antigen type A3; decreased phosphocreatine peak area after photic stimulation; and homozygotes for the short variant of the serotonin transporter gene. Most of the possible biological predictors of better lithium response, such as lower IMPase mRNA levels, white matter hyperintensity, lower brain intracellular pH, enhanced calcium response, and PLCG1-5 repeat had been detected as risk factors for bipolar disorder, suggesting that bipolar disorder responding well to maintenance lithium treatment is a distinct category having a certain neurobiological basis, although these findings need further replication. The search for biological predictors of lithium response is still in its infancy. Most of the laboratory or neuroimaging techniques used in these studies are not easily performed in clinical settings, so the development of an easy and useful laboratory test is needed.     

Chronic carbamazepine selectively downregulates cytosolic phospholipase A2 expression and cyclooxygenase activity in rat brain.

BACKGROUND: Carbamazepine is a mood stabilizer used as monotherapy or as an adjunct to lithium in the treatment of acute mania or the prophylaxis of bipolar disorder. Based on evidence that lithium and valproate, other mood stabilizers, reduce brain arachidonic acid turnover and its conversion via cyclooxygenase to prostaglandin E(2) in rat brain, one possibility is that carbamazepine also targets the arachidonic acid cascade. METHODS: To test this hypothesis, carbamazepine was administered to rats by intraperitoneal injection at a daily dose of 25 mg/kg for 30 days. RESULTS: Carbamazepine decreased brain phospholipase A(2) activity and cytosolic phospholipase A(2) protein and messenger RNA levels without changing significantly protein and activity levels of calcium-independent phospholipase A(2) or secretory phospholipase A(2). Cyclooxygenase activity was decreased in carbamazepine-treated rats without any change in cyclooxygenase-1 or cyclooxygenase-2 protein levels. Brain prostaglandin E(2) concentration also was reduced. The protein levels of other arachidonic acid metabolizing enzymes, 5-lipoxygenase and cytochrome P450 epoxygenase, were not significantly changed nor was the brain concentration of the 5-lipoxygenase product leukotriene B(4). CONCLUSIONS: Carbamazepine downregulates cytosolic phospholipase A(2)-mediated release of arachidonic acid and its subsequent conversion to prostaglandin E(2) by cyclooxygenase. These effects may contribute to its therapeutic actions in bipolar disorder.     

Metabonomic analysis identifies molecular changes associated with the pathophysiology and drug treatment of bipolar disorder

Bipolar affective disorder is a severe and debilitating psychiatric condition characterized by the alternating mood states of mania and depression. Both the molecular pathophysiology of the disorder and the mechanism of action of the mainstays of its treatment remain largely unknown. Here, (1)H NMR spectroscopy-based metabonomic analysis was performed to identify molecular changes in post-mortem brain tissue (dorsolateral prefrontal cortex) of patients with a history of bipolar disorder. The observed changes were then compared to metabolic alterations identified in rat brain following chronic oral treatment with either lithium or valproate. This is the first study to use (1)H NMR spectroscopy to study post-mortem bipolar human brain tissue, and it is the first to compare changes in disease brain with changes induced in rat brain following mood stabilizer treatment. Several metabolites were found to be concordantly altered in both the animal and human tissues. Glutamate levels were increased in post-mortem bipolar brain, while the glutamate/glutamine ratio was decreased following valproate treatment, and gamma-aminobutyric acid levels were increased after lithium treatment, suggesting that the balance of excitatory/inhibitory neurotransmission is central to the disorder. Both creatine and myo-inositol were increased in the post-mortem brain but depleted with the medications. Lastly, the level of N-acetyl aspartate, a clinically important metabolic marker of neuronal viability, was found to be unchanged following chronic mood stabilizer treatment. These findings promise to provide new insight into the pathophysiology of bipolar disorder and may be used to direct research into novel therapeutic strategies.     

Inositol as an add-on treatment for bipolar depression

Objective: Inositol is a constituent of the intracellular phosphatidyl inositol (PI) second messenger system, which is linked to various neurotransmitter receptors. Inositol crosses the blood–brain barrier in pharmacological doses, and has shown efficacy in a small double-blind study of unipolar depression. This pilot study evaluated its potential efficacy and safety in bipolar depression.

Methods: Twenty-four consenting adult men and women with DSM-IV bipolar depression (bipolar I=21; bipolar II=3) were randomly assigned to receive either 12 g of inositol or d -glucose as placebo for 6 weeks. Efficacy and safety ratings were done weekly. Thymoleptic medications (lithium, valproate, carbamazepine) in stable doses and at therapeutic levels at study entry were continued unchanged.

Results: Two subjects receiving placebo dropped out early due to worsening or non-adherence to the protocol. Among the 22 subjects who completed the trial, six (50%) of the inositol-treated subjects responded with a 50% or greater decrease in the baseline Hamilton Depression Rating Scale (HAM-D) score and a Clinical Global Improvement (CGI) scale score change of 'much' or 'very much' improved, as compared to three (30%) subjects assigned to placebo, a statistically non-significant difference. On the Montgomery–Asberg Depression Rating Scale (MADRS), eight (67%) of twelve inositol-treated subjects had a 50% or greater decrease in the baseline MADRS scores compared to four (33%) of twelve subjects assigned to placebo (p=0.10). Inositol was well tolerated with minimal side effects, and thymoleptic blood levels were unaltered.

Conclusions: These pilot data suggest a controlled study with an adequate sample size, and the appropriate rating scale may demonstrate efficacy for inositol in bipolar depression. The tolerability and the 'natural substance' aspect of inositol may be particularly appealing to subjects with bipolar depression.     

Chronic lithium administration alters a prominent PKC substrate in rat hippocampus

The therapeutic effect of lithium in the treatment of bipolar disorder exhibits a significant delay in the onset of action and a persistence of efficacy beyond abrupt discontinuation of treatment. Lithium is known to alter receptor-coupled phosphoinositide second messenger pathway in brain, resulting in indirect changes in an endogenous activator of protein kinase C (PKC). Such evidence has suggested that PKC may be involved in the mechanism of action of lithium in the brain. PKC represents a site wherein long-term regulatory changes in cell function occur through the phosphorylation of specific phosphoproteins involved in processes including neurotransmitter release and receptor activation. In studies of rats exposed to lithium, however, we have found no significant effects of chronic administration on the relative activity, subcellular distribution, or activation of PKC in hippocampus. We did find a major reduction in the in vitro PKC mediated phosphorylation of two major substrates, 83 kDa and 45 kDa, in hippocampus of rats exposed to chronic lithium and maintaining clinically relevant therapeutic levels in brain. Using immunoblot analysis we have identified a known myristoylated alanine-rich C kinase substrate (MARCKS) at 83 kDa. In vivo levels of MARCKS in hippocampus were found to be significantly reduced after chronic lithium exposure. These findings persist in animals withdrawn from lithium, but are not apparent following acute treatment. In light of the potential role of PKC substrates such as MARCKS in signal transduction and the fact that there appear to be changes in the intracellular concentration of MARCKS that parallel the time course of clinical action of lithium, we suggest the possible involvement of these proteins in its mechanism of action in the treatment of bipolar disorder.     

Brief chronic effects of lithium administration on rat brain phosphoinositides and phospholipids.

Lithium is known to exert its biochemical action on cells and tissues by inhibiting the enzymic conversion of inositol monophosphates to inositol. However, it is not clear whether this inhibitory action may lead to changes in the de novo biosynthesis of phosphatidylinositol and its phosphorylated derivatives. This biosynthetic scheme may have an important bearing with regard to the receptor-mediated signal transduction mechanism involving hydrolysis of polyphosphoinositides and release of inositol trisphosphate as second messenger for mobilization of intracellular calcium. In this study, the effects of brief chronic lithium administration on metabolism of brain phosphoinositides and other phospholipids were examined using the radiotracer technique with 32Pi as precursor. Sprague Dawley rats that were treated with lithium (3-4 meq/kg body wt) twice daily for 2-6 days consistently indicated an increase in the labeling of phosphatidylinositol 4,5-bisphosphates and a decrease in labeling of phosphatidylinositols and phosphatidylethanolamines. These phospholipid changes were found in both cortex and hippocampus and appeared to occur primarily in the synaptosomal fraction. Although the extent of the phospholipid changes could vary depending on both duration and dose levels of the lithium administered, these results demonstrated subtle effects of lithium on depressing the biosynthesis of phosphatidylinositol as well as phosphatidylethanolamine but perhaps a compensative increase in the synthesis of the phosphatidylinositol 4,5-bisphosphates.