Radioligand binding studies were done to investigate the effect of chronic administration of Amitriptyline on alpha1-adrenoceptor (alpha1-AR) receptor mediated response to inositol triphosphate (IP3) in rat brain. Our studies revealed a significant decrease in the densities of alpha1-ARs in cortex and cerebellum of rat brain after chronic administration of Amitriptyline (10 mg kg-1 b.wt.). However, there was no significant change in the affinity of [3H]prazosin to alpha1-ARs. Displacement studies showed that Amitriptyline has higher affinity for alpha1-AR with a Ki value of 182+/-16 nM. Significant change was observed in basal IP3 activity in cortex and cerebellum after Amitriptyline exposure. In cortex and cerebellum of experimental rats the NE (Norepinephrine) stimulated IP3 activity was significantly decreased (1460+/-102 DPM/g tissue; p<0.0001; 1188+/-112 DPM/g tissue; p<0.0001), when compared to NE stimulated IP3 activity (4152+/-286 and 3952+/-245 DPM/g tissue, respectively) in control rats. The decrease in NE stimulated IP3 activity in both regions may be due to the significant downregulation of alpha1-ARs in cortex after Amitriptyline exposure as these sites are positively coupled to IP3. The observed significant decrease in alphal-ARs with concomitant decrease in NE stimulated IP3 activity, after Amitriptyline treatment, suggests that Amitriptyline which has high affinity for these sites, acts by modulating the alpha1-AR receptor mediated response in brain.

The aim of the present study was to examine the effect of chronic antidepressants treatment on the density of α1-adrenoceptor (AR) subtypes in rat brain. Density of total α1 and α1A- and α1Β-ARs was measured in cortex and cerebellum of rats treated with amitriptyline (AMI), desipramine (DMI) and fluoxetine (FLX), (10 mg/kg body wt), for 30 days, using [3H]prazosin in presence and absence of WB-4101. The density of cortical total α1-ARs was significantly decreased with AMI (54%) and DMI (25%) treatment, without altering the affinity of the receptor. Fluoxetine did not alter the density of cortical α1-ARs. The density of cortical α1A-ARs was also significantly decreased with AMI (85%) and DMI (50%) treatment, without affecting the affinity. The density of cerebellar total α1-ARs was significantly decreased with AMI (37%), DMI (50%) and FLX (70%) treatment, without affecting the affinity for [3H]prazosin. The density of α1A-ARs was significantly decreased with AMI (67%), DMI (59%) and FLX (92%) treatment. α1B-AR density was decreased only with FLX (47%) and DMI (47%) treatment. Correspondingly the basal IP3 and NE (10 μM) stimulated IP3 levels were significantly decreased in AMI (47%), DMI (22%) and FLX (48%) treated rat cortex. The results suggest that chronic antidepressant (AD) treatment down-regulates the cortical and cerebellar total α1-ARs in rat brain. However, α1A subtype is predominantly down-regulated by AMI and DMI, where as FLX affects cerebellar α1A-ARs. The region-specific and subtype specific down-regulation of α1-ARs density, which occurs after prolonged AD treatment, may underline the therapeutic mechanism of action.

The discovery of cholinergic deficit in Alzheimer's disease (AD) patient's brain has triggered research efforts, using cholinomimetic approaches for their efficacy in AD therapy. Various therapies may be of potential clinical use in AD. Among these are cholinergic agents, which include muscarinic agonists, acetylcholinesterase inhibitors, and acetylcholine releasing agents. One of the muscarinic agonists tested in AD is arecoline and its bioisosters, which are widely explored as muscarinic receptor 1 agonist (M1 receptor agonist) in AD research. In this regard, five-membered heterocyclic ring system attached arecoline basic nucleus (N-methyl tetrahydropyridines) at third position has been extensively researched on. The present research involved synthesis of arecoline thiazolidinones 5(a–j) by using dipolar addition of 3-aminopyridine and alkyl/aryl carboxaldehydes in presence of gamma ferrite as catalyst. The resulting products were methylated and reduced to get desired products. Subsequently the synthesized arecoline thiazolidinones were subjected to in vitro muscarinic receptor binding studies using male Wistar rat brain (cerebral cortex) membrane homogenate and extended this in vitro study to in vivo pharmacological evaluation of memory and learning in male Wistar rats. Four derivatives (5a–5c and 5e) showed considerable M1 receptor binding affinity (in vitro) and elicited beneficial effects in vivo memory and learning models (Rodent memory evaluation, plus and Y maze studies).

A series of novel, potent and selective muscarinic receptor 1 agonists (M1 receptor agonists) that employ a key N -substituted morpholine arecoline moiety has been synthesized as part of research effort for the therapy of Alzheimer’s diseases. The ester group of arecoline (which is reported as mucarinic agonist) has been replaced by N- substituted morpholine ring. The structure activity relationship reveals that the increase in lipophilic carbon chain on the nitrogen atom of the morpholine ring increases the affinity of M1 receptor. In the present study, we are reporting N -amino acid substituted 9 ( a – k ) and dipeptides substituted 10 ( a – j ) and 11 ( a – j ) morpholino arecoline derivatives, along with their in vitro muscarinic binding studies by using [ 3 H]QNB and also in vivo evaluation of memory and learning in male Wistar rats (passive avoidance test plus maze studies) as M1 receptor agonist. Some molecules from the dipeptide series ( 10b , 10c and 10j ) showed potent M1 receptor agonist activity. Other derivatives also showed considerable M1 receptor binding affinity.

BACKGROUNDRecent studies have confirmed that alterations in the isoenzyme of pyruvate kinase (PK) provide tumor cells with selective growth advantage.AIMSOur aim was to establish the mean activity of the enzyme PK in human astrocytomas and to look for any trends in the activity with relation to histological grade.MATERIALS AND METHODSThe PK (EC 2.7.1.40) activity was measured in the tumor homogenate by spectrophotometric rate determination. DeltaAbsorbance at 340 nm (A 340 nm ) per minute was obtained using the maximal linear rate for both the test and the blank. Enzyme activity was estimated in the presence and absence of amino acid alanine.RESULTSThe mean PK level in astrocytomas was 3.5 +/- 2.0 mmol/min/mg protein, which was significantly higher (24%; P < 0.001) when compared to 2.8 +/- 0.3 mmol/min/mg protein in control brain. Highest PK activity was noted in grade 2 astrocytomas. In controls there was no change in PK activity in the presence of alanine. In grade 2 astrocytomas there was 7% decrease in mean PK activity in the presence of alanine, this difference in grade 3 astrocytomas was 33% and in grade 4 astrocytomas it was 61%. As the tumors were becoming malignant there was a graded increase in the levels of PK inhibition.CONCLUSIONSMean PK activity was significantly higher in astrocytomas. There was a graded increase in level of PK inhibition as the tumors were becoming more malignant.

Recent studies have conÞ rmed that alterations in the isoenzyme of pyruvate kinase (PK) provide tumor cells with selective growth advantage.[1] There are at least three isozymes of mammalian PK and in primary human brain tumors a shift occurs in the synthesis of M-type toward K-type. Isoenzyme shift can be documented with alanine-inhibition test as proposed initially by Van Veelen.[2] Most of the previous studies were concerned regarding PK inhibition. In this paper not only did we study PK inhibition but we also established the mean PK activity in astrocytomas.

Earlier we have reported the effect of arecoline thiazolidinone and morpholino arecoline analogues as muscarinic receptor 1 agonist in Alzheimer's dementia models. To elucidate further our SAR study on the chemistry and muscarinic receptor binding efficacy, a series of novel N-alkyl/aryl substituted thiazolidinone arecoline analogues 6(a–m) were designed and synthesized from 3-pyridine carboxaldehyde by reacting with different amines in the presence of γ-ferrite as catalyst and subjected to in vitro muscarinic receptor binding studies using male Wistar rat brain membrane homogenate and extended to in vivo pharmacological evaluation of memory and learning in male Wistar rats. Derivative 6j having diphenylamine moiety attached to nitrogen of thiazolidinone showed significant affinity for the M1 receptor binding.

EDITOR: Phenytoin is generally prescribed to patients with supratentorial tumours to decrease the risk of seizures. Earlier studies showed that plasma phenytoin concentration may not be in the therapeutic range despite continued therapy [1]. During craniotomy for a supratentorial tumour, an intraoperative loading dose of phenytoin is generally used to prevent postoperative seizures [2,3]. In our institution, it has been common practice not to administer phenytoin intraoperatively. To understand the consequences of our practice of withholding intraoperative phenytoin, we measured perioperative serum phenytoin concentration in a group of patients undergoing supratentorial tumour surgery. We also tried to determine the factors influencing postoperative serum phenytoin concentrations. Twenty-five adult patients (ASA I or II) of either sex, receiving phenytoin for a period not less than 7 days before supratentorial surgery, were studied after institutional approval and informed consent. On the day of surgery, 300 mg of phenytoin was administered either orally or intravenously 4 h before surgery. The anaesthetic technique comprised of induction with thiopentone (5–6 mg kg−1), tracheal intubation facilitated by a muscle relaxant and maintenance with either isoflurane or propofol. Intraoperative analgesia was provided by fentanyl. Serum phenytoin concentration was measured before induction, immediately after surgery and 24 h after surgery. The assay, performed by a chemiluminescence technique using an Immulite Assay Kit® (DPC®; Los Angeles, CA, USA), permitted the measurement of total phenytoin concentration. The following parameters were recorded in all patients: duration of anaesthesia and surgery, volume of crystalloids, colloids and blood products administered, volume of urine output and blood loss, and occurrence of immediate postoperative seizures. A repeated-measures analysis of variance (ANOVA) with Bonferroni's test was used to find out significant differences among the preinduction, immediate postoperative and delayed postoperative serum phenytoin concentrations. Study variables in patients with therapeutic and subtherapeutic concentrations of phenytoin were compared by one-way ANOVA for continuous data and a χ2-test for categorical variables. Pearson's test was used to correlate preinduction phenytoin concentration and its decrease in the immediate postoperative period. Logistic regression analysis was used to determine the independent predictors of immediate postoperative subtherapeutic serum phenytoin concentration. A P value of <0.05 was considered significant. There were 17 male and 8 female patients in the study. Their age was 38 ± 12 yr and body weight was 56 ± 11 kg. Twelve patients had a preoperative history of seizures. The preinduction serum phenytoin concentration was highly variable among the patients (range 2.5–37.3 μg mL−1 (95% CI = 9.8–17.8 μg mL−1)). Despite continuous medication until the morning of surgery, 11 patients (44%) had a subtherapeutic concentration of serum phenytoin (normal range 10–20 μg mL−1) in the preinduction sample. Serum phenytoin concentration was significantly lower in the immediate postoperative sample compared with the preinduction sample (9.5 ± 7.0 vs. 13.8 ± 9.4 μg mL−1; P < 0.001). The concentration increased significantly in the delayed postoperative sample (11.8 ± 8.0 μg mL−1; P < 0.001). The decrease in phenytoin concentration in the immediate postoperative sample correlated with its preinduction value (P < 0.01; r = 0.8). Seizures within 24 h of surgery occurred in two patients. Only one of these patients had a preoperative history of seizures. Serum phenytoin concentration was within the therapeutic range (16.3 and 10.4 μg mL−1) in both the patients. In the immediate postoperative period, serum phenytoin concentration was in a subtherapeutic range in 15 out of the 25 study patients. On univariate analysis, the variables that were significantly different between the therapeutic and subtherapeutic groups were the patient's gender, preinduction phenytoin concentration, blood loss, blood transfusion, duration of surgery and duration of anaesthesia (Table 1). Of these, preoperative phenytoin level, intraoperative blood transfusion and the duration of surgery/anaesthesia were found to be the independent predictors of low serum phenytoin concentration in the immediate postoperative period (P < 0.05).Table 1: Study variables in patients with subtherapeutic and therapeutic postoperative phenytoin concentration.The value of prophylactic administration of antiepileptic drugs in patients with brain tumours remains controversial. Some authors claim a significant decrease in the incidence of seizures in the postoperative period [2,3] with preoperative antiepileptic therapy. However, in one study, phenytoin doses aimed at maintaining serum phenytoin concentrations in the 10–20 μg mL−1 range did not decrease the incidence of postoperative seizures [4]. One meta-analysis showed a statistically insignificant reduction of postoperative convulsions with prophylactic anticonvulsant and suggested the need for further investigation of the issue [5]. In our study, preoperative serum phenytoin concentration was highly variable with 44% of the patients having subtherapeutic concentrations as has been reported earlier [1]. Widely variable clearance, as is known to occur even in normal individuals, could probably be the reason for this variation. Another possible cause is the interaction between dexamethasone and phenytoin as has been reported earlier [6]; all our patients had been receiving dexamethasone for several days before surgery. There was a linear correlation between preoperative serum phenytoin concentration and its decrease in the intraoperative period, which is possibly related to the first-order kinetics of phenytoin. We found that preinduction serum phenytoin concentration, need for blood transfusion and the duration of surgery/anaesthesia were independent predictors of low phenytoin concentration in the immediate postoperative period. A simple process of dilution caused by blood and fluid replacement might be responsible for the decrease in phenytoin concentration. Long-duration surgery might have caused increased excretion and lower serum concentration. In a recent study by Yeh and colleagues [1], less than 50% of the patients had a therapeutic level of serum phenytoin and the predictors of low serum phenytoin concentration were the same as in our study. Despite withholding the intraoperative phenytoin dose, only two patients had postoperative seizures within 24 h. The small sample size in the study prevents drawing serious conclusions regarding the incidence of perioperative seizures with and without intraoperative loading. Given the subtherapeutic phenytoin concentration in a major proportion of our patients, it is advisable to administer an intraoperative dose of phenytoin to achieve the therapeutic level. Whether these additional doses cause toxic levels of phenytoin in patients in whom the metabolic pathways are already saturated also remains to be seen. A more rational approach would be to decide the dosing based on serum phenytoin levels measured preoperatively. It would also be interesting to study the influence of perioperative corticosteroids on serum phenytoin levels.

Amine transporters are major target for development of various pharmacological agents to treat behavioral disorders. Serotonin transporters (SERT) have been implicated in the etiology of depression and drugs acting on SERT can be effective in treating depression. The aim of the present study was to study the in vivo effect of various antidepressants on [ 3 H]paroxetine binding to SERT in regions of rat brain. Rats were treated with tricyclic antidepressant (TCAs) such as amitriptyline (AMI), serotonin/norepinephrine reuptake inhibitor (SNRIs) such as clomipramine (CMI), and selective serotonin reuptake inhibitors (SSRIs) such as fluoxetine (FLX) and citalopram (CIT) (10 mg/kg body wt.) for 30 days. Density of SERT was measured in cortex and hippocampus using [ 3 H]paroxetine (0.03–1.0 nM) in presence and absence of 10 μM fluoxetine as displacer. It was observed that the density of cortical SERT was significantly decreased with CMI (68%, P < 0.0001), FLX (67%, P < 0.0001), CIT (54%, P < 0.0001), and AMI (52%, P < 0.0001) treatment, when compared to the density of 120.7 ± 4.0 fmol/mg protein in control rats, without altering the affinity (Kd) of [ 3 H]paroxetine to the transporters. The density of SERT in hippocampus was also significantly decreased with FLX (65%, P < 0.0001), CMI (54%, P < 0.0001), CIT (52%, P < 0.0001) and AMI (46%, P < 0.0001) treatment, when compared to the density of 74.0 ± 2.6 fmol/mg protein in control rats, without altering the affinity of [ 3 H]paroxetine to the transporters. Displacement study showed high affinity for CMI > CIT > FLX. The results suggest that chronic antidepressant treatment significantly down-regulates both cortical and hippocampal SERT in rat brain and SSRIs have high affinity for SERT than TCAs.

As part of our continuing effort aimed at the development of selective, efficacious and centrally active M1 muscarinic agonists for the treatment of Alzheimer's presenile dementia, a series of N-arylthioureas substituted 3-morpholino arecoline derivatives 9(a-j) were synthesized by using N-benzyl amino ethanol coupling with alpha-bromo acetyl pyridine followed by reduction and cyclization to develop a new class of M1 receptor agonists. Subsequently the synthesized compounds were subjected to in vitro radioligand M1 receptor affinity studies, IP3 formation studies and also to in vivo pharmacological evaluation of memory and learning in male Wistar rats. Derivatives with chloro (9f) and methoxy (9c) groups on the para position of the benzene ring attached to the nitrogen of thiourea showed several fold high affinity for the M1 receptor (in vitro) among all the synthesized molecules 9(a-j), and also significantly elevated IP3 levels and as well elicited beneficial effects in vivo in memory and learning models in rats (rodent memory evaluation, plus and Y maze studies).

A series of novel, potent, and selective muscarinic receptor 1 agonists (M1 receptor agonists) that employ a key N-substituted morpholine Arecoline moiety has been synthesized as part of research effort for the therapy of Alzheimer’s diseases. The ester group of arecoline (which is reported as muscarinic agonist) has been replaced by N-substituted morpholine ring. The structure–activity relationship reveals that the electron donating 4-substituted sulfonyl derivatives (9a, 9b, 9c, and 9e) on the nitrogen atom of the morpholine ring increases the affinity of M1 receptor binding 50- to 80-fold greater than the corresponding arecoline. Other derivatives also showed considerable M1 receptor binding affinity.

The genetic basis of bipolar disorder (BPD) and schizophrenia (SCZ) has been established through numerous clinical and molecular studies. Although often considered separate nosological entities, evidence now suggests that the two syndromes may share some genetic liability. Recent studies have used a composite phenotype (psychosis) that includes BPD, SCZ, psychosis not otherwise specified, and schizoaffective disorder, to identify shared susceptibility loci. Several chromosomal regions are reported to be shared between these syndromes (18p, 6q, 10p, 13q, 22q). As a part of our endeavor to scan these regions, we report a positive linkage and association finding at 18p11.2 for psychosis. Two-point linkage analysis performed on a series of 52 multiplex pedigrees with 23 polymorphic markers yielded a LOD score of 2.02 at D18S37. An independent set of 159 parent offspring trios was used to confirm this suggestive finding. The TDT analysis yielded support for association between the marker D18S453 and the disease allele (chi2 = 4.829, P < 0.028). This region has been implicated by several studies on BPD [Sjoholt et al. (2004); Mol Psychiatry 9(6):621-629; Washizuka et al. (2004); Biol Psychiatry 56(7):483-489; Pickard et al. (2005); Psychiatr Genet 15(1):37-44], SCZ [Kikuchi et al. (2003); J Med Dent Sci 50(3):225-229; Babovic-Vuksanovic et al. (2004); Am J Med Genet 124(3):318-322] and also as a shared region between the two diseases [Ishiguro et al. (2001); J Neural Transm 108(7):849-854; Reyes et al. (2002); Mol Psychiatry 7(4):337-339; Craddock et al. (2005); J Med Genet 42(3):193-204]. Our findings provide an independent validation of the above reports, and suggest the presence of susceptibility loci for psychoses in this region.

OBJECTIVE Lithium remains the most widely used treatment for bipolar disorder; however, the molecular mechanisms underlying its therapeutic actions have not been fully elucidated. We studied the in-vivo effect of lithium on the density of alpha-adrenoceptor (alpha-AR) and beta-AR subtypes and linked second messenger systems in the rat brain. METHODS The densities of alpha(1)-ARs, alpha(2)-ARs, and beta(1)-ARs and beta(2)-ARs in the cortex and cerebellum of rats treated with lithium (0.4%), orally, for 30 days were measured using [(3)H]prazosin, [(3)H]clonidine and [(3)H]CGP-12177, respectively. The activity of adenylyl cyclase (AC) and levels of inositol trisphosphate (IP3), both second messengers linked to these receptors, were estimated using [(3)H]ATP and [(3)H]myoinositol, respectively. RESULTS A significant decrease in the densities of cortical alpha(1)-ARs (85%, p < 0.0001), alpha(2)-ARs (50%, p < 0.0001), beta(1)-ARs (26%, p < 0.0001) and beta(2)-ARs (25%, p < 0.0001) was observed after lithium treatment. However, only the density of alpha(1)-ARs was significantly decreased (25%, p < 0.0001) in the cerebellum. The affinity of [(3)H]prazosin for cerebellar alpha(1)-ARs was increased. A small, but statistically significant, increase (19%, p < 0.0001) in the density of total beta-ARs was seen in the cerebellum, without altering the affinity of the radioligand for these receptors. Basal AC activity was not altered in the lithium-treated rat cortex. However, the norepinephrine-stimulated AC activity, which represents alpha(2)-AR-linked and beta-AR-linked AC, was significantly increased (66%, p < 0.0001). Both basal IP3 formation and norepinephrine-stimulated IP3, which represents alpha(1)-AR-linked phospholipase C activity, were significantly decreased (50%, p < 0.0001) in the lithium-treated rat cortex. CONCLUSION Our results suggest that long-term administration of lithium treatment downregulates the cortical, but not cerebellar, alpha(1)-ARs, alpha(2)-ARs, beta(1)-ARs and beta(2)-ARs. Thus, it may be concluded that lithium induces region-specific and differential functional downregulation of alpha-AR and beta-AR subtypes in the rat brain.

It has been well established that alterations in biogenic amine receptor functions are involved in the pathophysiology of behavioral disorders. Serotonin receptor mediated response has been implicated in the mechanism of action of various antidepressants (ADs). However, the molecular mechanisms by which antidepressant drugs bind to and regulate 5-HT receptors remain an unsolved problem. In this study, the in vivo effect of chronic administration of imipramine (5 mg/kg body weight), a TCA, and fluoxetine (10 mg/kg body weight), a SSRI, on 5-HT2 receptor-mediated second messenger system is studied in Sprague–Dawley rats. Both imipramine and fluoxetine treatment significantly down regulated (38%) 5-HT2 receptor density in cortex, without changing the affinity of [H]spiperone. A significant decrease in 5-HT stimulated IP3 formation, from [ H] myoinositol, in cortical slices was seen with imipramine (48%) and fluoxetine (58%) treatment. 5-HT sensitive intracellular Ca mobilization in cortical membranes, as measured by FURA-2AM, was significantly decreased with fluoxetine (60%) and imipramine (50%) treatment. These findings were correlated with findings in human autopsy brain samples from MDP patients. This study shows that the mode of action of these ADs, apart from being uptake inhibitors, is through 5-HT2 receptor-mediated IP3 formation, which in turn modulates intracellular Ca levels in brain. These ADs reduce the hypersensitivity of 5-HT2 receptor-mediated response, which is observed in depressive patients, thus relieving depression.

The aim of the present study was to examine the effect of chronic tricyclic antidepressants (TCAs) treatment on the density of α-adrenoceptors in the rat brain. Density of α1- and α2-adrenoceptors was measured in cortex and hippocampus of rats treated with imipramine (IMI, 5mg/kg body weight), desipramine (DMI, 10mg/kg body weight), clomipramine (CMI, 10mg/kg body weight) and amitriptyline (AMI, 10mg/kg body weight), for 40 days, using [3H]prazosin and [3H]clonidine, respectively. The density of cortical α1-adrenoceptors was significantly decreased with IMI (46%), DMI (21%), CMI (50%) and AMI (67%) treatment, without altering the affinity of the receptor. The density of cortical α2-adrenoceptors was also significantly decreased with DMI (69%), CMI (81%) and AMI (80%) treatment, without affecting the affinity for [3H]clonidine. The density of hippocampal α1-adrenoceptors was significantly decreased only with AMI treatment (47%), without affecting the affinity for [3H]prazosin. However, no change in hippocampal α2-adrenoceptor density was observed with any of these TCAs. The results suggest that chronic antidepressant (AD) treatment downregulates the cortical, but not hippocampal, α1- and α2-adrenoceptors in rat brain. The region-specific downregulation of α1- and α2-adrenoceptors density, which occur after prolonged AD treatment, may underline the therapeutic mechanism of action.