High Doses Of Colchicine Research Articles

Low-Dose Colchicine Ameliorates Doxorubicin Cardiotoxicity Via Promoting Autolysosome Degradation.

The only clinically approved drug that reduces doxorubicin cardiotoxicity is dexrazoxane, but its application is limited due to the risk of secondary malignancies. So, exploring alternative effective molecules to attenuate its cardiotoxicity is crucial. Colchicine is a safe and well-tolerated drug that helps reduce the production of reactive oxygen species. High doses of colchicine have been reported to block the fusion of autophagosomes and lysosomes in cancer cells. However, the impact of colchicine on the autophagy activity within cardiomyocytes remains inadequately elucidated. Recent studies have highlighted the beneficial effects of colchicine on patients with pericarditis, postprocedural atrial fibrillation, and coronary artery disease. It remains ambiguous how colchicine regulates autophagic flux in doxorubicin-induced heart failure. Doxorubicin was administered to establish models of heart failure both invivo and invitro. Prior studies have reported that doxorubicin impeded the breakdown of autophagic vacuoles, resulting in damaged mitochondria and the accumulation of reactive oxygen species. Following the administration of a low dose of colchicine (0.1 mg/kg, daily), significant improvements were observed in heart function (left ventricular ejection fraction: doxorubicin group versus treatment group=43.75%±3.614% versus 57.07%±2.968%, P=0.0373). In terms of mechanism, a low dose of colchicine facilitated the degradation of autolysosomes, thereby mitigating doxorubicin-induced cardiotoxicity. Our research has shown that a low dose of colchicine is pivotal in restoring the autophagy activity, thereby attenuating the cardiotoxicity induced by doxorubicin. Consequently, colchicine emerges as a promising therapeutic candidate to improve doxorubicin cardiotoxicity.

Effect of increasing doses of colchicine on the treatment of 333 COVID-19 inpatients.

Previous research done in Bulgaria demonstrated a fivefold reduction in mortality from COVID-19 with increased doses of colchicine from two hospitals in the country. We report here a further 333 cases of COVID-19 inpatients, treated with different doses of colchicine and its effect on mortality. A case-control comparison from two additional hospitals was conducted between increased doses of colchicine and added bromhexine to standard of care (SOC) versus current SOC. Risk and odds ratio, as well as subgroup analysis, was conducted with newly reported data, alongside aggregate data from all hospital centers to determine the extent of mortality reduction in COVID-19 inpatients. There was a clear reduction in the mortality of inpatients with increasing doses of colchicine-between twofold and sevenfold. Colchicine loading doses of 4 mg are more effective than those with 2 mg. Despite these doses being higher than the so-called "standard doses," colchicine inpatients experienced lower mortality than SOC patients (5.7% vs. 19.53%). This mortality benefit was evident in different age subgroups, with a 4-mg loading dose of colchicine proving slightly superior to a 2-mg loading dose. Colchicine led to an overall relative risk reduction of 70.7%, with SOC patients having 3.91 higher odds of death. The safety of the doses was not different than the reported in the summary of product characteristics. Inpatients in Bulgaria with added colchicine and bromhexine to SOC achieved better clinical and mortality outcomes than those on SOC alone. These results question the World Health Organization-recommended strategy to inhibit viral replication. We posit that our treatment strategy to inhibit the Severe acute respiratory syndrome coronavirus 2 entry into the cell with inhaled bromhexine and the hyperactivated NLRP3 inflammasome with higher doses of colchicine, prevents the development of cytokine storm. The timing of the initiation of treatment seems critical.

Comparison of treatment of COVID-19 with inhaled bromhexine, higher doses of colchicine and hymecromone with WHO-recommended paxlovid, molnupiravir, remdesivir, anti-IL-6 receptor antibodies and baricitinib

Millions of publications and thousands of clinical trials have not led to the discovery of an effective treatment for COVID-19. We believe that the reason for this is the inaccurate strategy of inhibiting target molecules involved in the pathogenesis of the disease. The leading cause of death in COVID-19 is the cytokine storm, which is caused by an NLRP3 inflammasome hyperreaction. WHO recommends for the outpatients treatment drugs blocking the replication of SARS-CoV-2. However, viral load and replication are not directly related to NLRP3 inflammasome hyperreactivity. This also explains the partial success of the WHO favorite paxlovid to reduce hospitalizations (51%). For hospital treatment, WHO suggests antibodies against the interleukin-6 receptor and Janus kinase (JAK) inhibition. Although important, IL-6 is one of dozens of cytokines elevated as a consequence of cytokine storm. The JAK inhibitor baricitinib inhibited the effect of not only IL-6 but also other elevated cytokines. But if the NLRP3 inflammasome is inhibited, the cytokines will not be elevated, and there will be no need for baricitinib. All medicines recommended by the WHO are distinguished by their very high prices. Our therapeutic strategy is based on inhibition of SARS-CoV-2 entry into the cell and inhibition of the NLRP3 inflammasome. We offer two readily available, cheap and well-known medications - bromhexine hydrochloride and colchicine. The many studies on the treatment of COVID-19 so far have not produced the expected result. The devil is buried in the details. For bromhexine, the reason is the way and its late application. Bromhexine is most effective when given prophylactically or started by inhalation after contact with a person with COVID-19. Its earliest possible application is crucial for its effect. Increased doses of colchicine are necessary for COVID-19 treatment due to the fact that it accumulates in leukocytes, and this leads to inhibition of NLRP3. The high doses we administer have been given widely in the past and are completely safe. Our highest dose is about 5 times lower per kg of weight than the lowest severe toxic dose of colchicine described. Our results show about a 5-fold decrease in hospital mortality and almost complete prevention of hospitalizations if outpatients are treated with inhaled bromhexine and colchicine.

Impact of Colchicine on Histology of Testis in Rats

Colchicine is a drug widely used for the management of many disorders, such as acute gout and Behçet’s disease. It is also prescribed for the treatment of pericarditis, atrial fibrillation coronary artery diseases, and secondary amyloidosis. In case this drug is used at the early stages of coronavirus infection, its anti-inflammatory properties may reduce the severe inflammatory reactions related to a cytokine storm by affecting the inflammasome. The purpose of the present study was to determine the toxicity of Colchicine on testis in rats from different age groups for 10 days. A total of 27 male Wistar rats were divided into three groups. The rats in group I (control group) were administered distilled water by oral gavage. Group II consisted of young rats (5-6 months old) who orally received Colchicine 3 mg/kg body weight. Group III entailed rats of 14-16 months who were orally administered colchicine 3 mg/kg body weight. The testis of the treated groups was dissected and examined for histological changes and morphometrical analysis. The obtained results indicated that high doses of Colchicine (3 mg/kg body weight) could induce tissue damage to the testis, including degeneration and necrosis of both Sertoli and Leydig cells with irregular divisions of germinal epithelium, even when it was used for short periods (10 days). In the elderly treated rats, there were severe tissue damages, including degeneration and necrosis of germinal epithelium with irregular divisions of germ cells, necrosis of Sertoli and Leydig cells with sloughing of germinal epithelium toward the lumen of the tubule. Therefore, there is a need to conduct more studies to investigate the side effect of Colchicine as it is excessively used in the management of coronavirus.

The expanded spectrum of arthritis in children with familial Mediterranean fever.

Familial Mediterranean fever (FMF) is the most common monogenic autoinflammatory disease that can present with various forms of arthritis. This retrospective study aims to evaluate the characteristics of patients with arthritis in a large pediatric cohort of FMF patients. The demographic and clinical data were extracted from electronic medical records. Patients with arthritis were grouped as arthritis of FMF and arthritis of FMF-associated diseases. A total of 541 patients were followed with a diagnosis of FMF in the last 5years. Acute arthritis of FMF (n: 138) was the most common cause. It showed a recurrent course in the majority with a longer duration than other attack symptoms. Significantly higher frequencies of biallelic exon 10 and M694V mutations, erysipelas-like erythema, and protracted febrile myalgia were detected in these patients, particularly in those older than 2years of age. Sacroiliitis of FMF was the second most common cause (n: 19). Patients with acute arthritis and sacroiliitis of FMF needed higher doses of colchicine. One patient with neonatal-onset FMF and M694V homozygosity was diagnosed with protracted arthritis. Arthritides of FMF-associated diseases including IgA vasculitis (n: 10), juvenile idiopathic arthritis (n: 9), chronic nonbacterial osteomyelitis (n: 5), and inflammatory bowel disease (n: 2) were detected in 26 patients. Arthritis is an important clinical finding of FMF mostly associated with M694V mutations. The frequency of protracted arthritis is declined, whereas sacroiliitis of FMF and arthritis of associated diseases expand the spectrum of arthritis. This study represents the changing face and current perspectives of arthritis in FMF. Key Points • Arthritis is an important clinical finding of familial Mediterranean fever (FMF) that can present in various forms • Arthritis is most likely associated with M694V mutations • The frequency of protracted arthritis is declined whereas sacroiliitis of FMF and arthritis of associated diseases expand the spectrum of arthritis in FMF.

Pharmacokinetics and Colchicine in Pediatric and Adult Patients with Familial Mediterranean Fever

This study sought to determine the appropriate starting dose of colchicine in children aged 2 to 4 years with familial Mediterranean fever (FMF) based on steady-state pharmacokinetics in pediatric patients with FMF from 2 to less than 16 years and adult patients with FMF from 16 to 65 years. Outpatients received colchicine for 90 days starting with a fixed dose for 14 days (blood sampling days 14 and 15). After starting doses of colchicine (0.6 mg/day [2 to less than 4 years], 0.9 mg/day [from 4 to less than 6 years], 0.9 mg/day [from 6 to less than 12 years], 1.2 mg/day [from 12 to less than 16 years], and 1.2 mg/day [from 16 to less than 65 years]), the observed steady-state pharmacokinetic parameters were comparable across age groups, despite the higher doses of colchicine on a mg/kg/day basis in the younger age groups. An exception occurred with once-daily colchicine, whereby mean Cmax for colchicine was higher in patients 4 to less than 6 years (9.4 ng/mL) compared with the younger and older age groups (6.1-6.7 ng/mL). Mean AUC0?24h values in children 2 to less than 4, 6 to less than 12, and 12 to less than 16 years were similar to those in adults. However, mean AUC0?24h values in children 4 to less than 6 years were 25 percent higher than those observed in adults. The results show that the recommended starting dose for children 2-4 years and 4-6 years should be 0.6 mg/day (half the US adult dose). Children aged 6 to less than 12 years should receive 0.9 mg/day (i.e. three-quarters of the US adult dose). The safety of colchicine in children 2 to less than 4 years was comparable to that in older children and adults.

Recent advances in management of gout

Incidence and prevalence of gout have markedly increased over the last few decades in keeping with the rise in prevalence of obesity and metabolic syndrome. Until recently, management of gout in patients with associated metabolic syndrome and comorbid illnesses such as renal impairment was difficult because of limited treatment options. However, significant progress has been made in the last few years, with introduction of new treatments such as interleukin-1 antagonists for management of acute gout, and febuxostat and pegloticase for chronic gout. The association of gout with alcohol, dietary purines and fructose ingestion has been confirmed in large prospective studies, thus enabling the clinician to now provide evidence-based advice to patients. Recent efficacy and safety data favour lower over higher doses of colchicine, and oral corticosteroids over non-steroidal anti-inflammatory drugs for patients with acute gout. Local ice therapy might help to differentiate gout from other forms of inflammatory arthritis, and supplementation with vitamin C help to reduce risk of gout. Several other drugs with rational mechanisms of action are in the pipeline, and likely to be introduced over the next few years. A new era has thus begun in the field of gout.

Fatal poisoning in children: Acute Colchicine intoxication and new treatment approaches

Background. Colchicine poisoning is potentially life-threatening. Deaths generally result from hypovolemic shock and cardiovascular collapse or secondary to rapidly progressive multiorgan failure. Objective. The purpose of this study is to discuss the clinical effects, treatments and outcomes of pediatric colchicine poisoning and highlight the possible benefits of urgent plasma and whole blood exchange therapy for those patients who were believed to ingest potentially lethal doses of the drug. Methods. Current study was designed as an observational case series study. The medical records of children aged 0–16 years who were hospitalized for colchicine poisoning at the Pediatric Intensive Care Unit of, between November 1985 and March 2011 were retrospectively evaluated. Results. We present twenty-three children with colchicine poisoning. Nausea and vomiting were the most common presenting complaint, in 70% of patients. Sixteen of the 23 cases presented after ingesting sub-toxic doses of colchicine (< 0.5 mg/kg), whereas 3 patients had consumed toxic doses of the drug (0.5–0.8 mg/kg). The remaining 4 patients were hospitalized after taking colchicine at a lethal dose (> 0.8 mg/kg). Three patients (13%) died. Conclusions. Any patient suspected of ingesting high doses of colchicine should prompt immediate fluid and electrolyte resuscitation and invasive hemodynamic monitorization in a pediatric intensive care unit. Although there is lack of strong evidence, early initiation of either whole blood or plasma exchange may be considered in patients presenting with lethal-dose colchicine intoxication. These reported experience of us put forth further research for consideration.

Disruption of axoplasmic transport induces mechanical sensitivity in intact rat C‐fibre nociceptor axons

Peripheral nerve inflammation can cause axons conducting through the inflamed site to become mechanically sensitive. Axonal mechanical sensitivity (AMS) of intact axons may explain symptoms in a diverse number of conditions characterized by radiating pain evoked by movements of the affected nerve. Because nerve inflammation also disrupts axoplasmic transport, we hypothesized that the disruption of axoplasmic transport by nerve inflammation could cause the cellular components responsible for mechanical transduction to accumulate and become inserted at the inflamed site, causing AMS. This was tested by examining AMS in C-fibre nociceptors following the application of axoplasmic transport blockers (colchicine and vinblastine) to the sciatic nerve. Both 10 mm colchicine and 0.1 mm vinblastine caused AMS to develop in 30.6% and 33.3% of intact axons, respectively (P < 0.05 compared to sham treatment). Since high doses of colchicine (> 50 mm) can damage axons, and inflammation is involved in the removal of axonal debris, experiments were performed to assess conduction across the treatment site as well as signs of inflammation. Results indicated minimal axonal loss (95% of A- and C-fibres conducting), consistent with the normal microscopic appearance of the colchicine treatment site and absence of ED1-positive (recruited) macrophages. In a separate series of experiments, the block of axoplasmic transport proximal to a localized neuritis significantly reduced inflammation-induced AMS (15.6% compared to 55.6%; P < 0.05), further supporting that the components necessary for AMS are moved by anterograde transport. In summary, nerve inflammation that causes the disruption of axoplasmic transport in patients with painful conditions may result in the accumulation and insertion of mechanosensitive elements at the inflamed site.

Colchicine in acute gout: the need for a reappraisal

Colchicine in acute gout: the need for a reappraisal

Colchicine treatment in children with familial Mediterranean fever

Prophylactic colchicine therapy has been shown to be a safe and effective method of eliminating the attacks and preventing the development of amyloidosis in patients with familial Mediterranean fever (FMF). However, information about effective dosages that control FMF attacks and prevent amyloidosis in childhood is not available. The aim of this study is to determine the 'effective colchicine dose' for children in terms of body weight and surface area. Sixty-two (34 male, 28 female) children with FMF were selected and colchicine treatment was initiated by giving 0.5-1 mg/day to each patient. The dose was gradually increased up to a maximum 2 mg/day in unresponsive patients; mean duration of therapy was 45.6 +/- 35.5 months. When the 'optimal effective dosage' (i.e. the one that reduced the frequency of attacks and ESR, CRP and fibrinogen levels during the attack-free period) was achieved, the optimal effective dose was calculated according to the body weight and body surface area for each patient. Based on these values 'mean colchicine dose' was computed for the study group and values for different age groups were evaluated. Mean colchicine doses according to the body weight and surface area of the whole group were found to be 0.03 +/- 0.02 mg/kg/day and 1.16 +/- 0.45 mg/m(2)/day, respectively. It was shown that children less than 5 years of age might need colchicine doses as high as 0.07 mg/kg/day or 1.9 mg/m(2)/day. These dosages are approximately 2.5-3 times more than the 'mean colchicine dose' for children aged 16-20 years. These results clearly show that small children need higher doses of colchicine in order to control their attacks. Thus, we conclude that colchicine, when given according to body weight or body surface area, would be more effective in childhood.

Kinetics of micronucleus induction and cytotoxic activity of colchicine in murine erythroblast in vivo

In previous studies, we inferred some pharmacokinetic and pharmacodynamic parameters of alkylating agents and antimetabolites by comparing their kinetics of micronucleated polychromatic erythrocyte (MN-PCE) induction with the one obtained after the exposure to gamma rays in peripheral blood of mice, assuming that radiation acts immediately because it does not require absorption and distribution in the organism. According to our earlier studies, the kinetics of MN-PCE induction depends mainly on the following: (i) the cytotoxic effects that in turn could affect the duration of cell division; (ii) the pharmacokinetics including the metabolic activation requirement; and (iii) the mechanism of MN induction. The aim of the present study was to analyze the kinetics of MN-PCE induction by an aneuploidogen that induces micronuclei by acting on the achromatic spindle. The kinetics of MN-PCE induction by colchicine, as well as the reduction in the PCE frequency over time was determined in peripheral blood of mice treated with different doses of the aneuploidogen. The genotoxic effect, established as the area beneath the curve (ABC) of MN-PCE versus time-response, indicates an almost directly proportional relationship with respect to dose. Similarly, the relationship between dose and cytotoxic effect determined as the ABC of PCE versus time was inversely proportional, suggesting a relationship between both endpoints and doses administered. However, the number of cells affected by these two phenomena indicates that cytotoxicity is not necessarily caused, or at least not only by genotoxicity. The analysis of the kinetics of MN-PCE induction after the treatment with non-cytotoxic dose of colchicine, indicates that the MN-PCE appear in the blood stream at almost the same time, as occurs after the exposure to gamma rays; in spite of the differences in the cell cycle stage in which they can cause micronucleus (MN). Perhaps the fact that cells are not synchronized does not permit one to observe some difference in the time they appear in the blood. These results suggest that colchicine acts rapidly after exposure. The elimination half-life of colchicine is 17 h, suggesting that colchicne is disposable for long time. With high doses of colchicine the pharmacokinetic parameters increases substantially. These data imply that low doses of colchicine are slightly cytotoxic, and that under this circumstances colchicines arrives rapidly to hemopoyetic tissues and acts for several hours.

Cardiotoxicity of colchicine in the rat.

Colchicine poisoning may be lethal and a decrease in cardiac function has been reported in several case reports, but the precise cardiotoxicity of colchicine remains unknown. The experimental in vitro study assessed the intrinsic contractility of left ventricular papillary muscle in rats, 24 h after administration of intraperitoneal colchicine or saline. The administration of colchicine (2 or 4 mg.kg-1) in adult Wistar rats markedly impaired intrinsic myocardial contractility, as shown by a decrease in maximum shortening velocity (-32 and -61%, respectively), active isometric force (-47 and -65%, respectively), and peak power output (-57 and -69%, respectively) of left ventricular papillary muscle. Colchicine impaired isotonic relaxation and load dependence of relaxation, suggesting a decrease in sarcoplasmic reticulum function. Conversely, colchicine significantly accelerated isometric relaxation, suggesting a decrease in calcium myofilament sensitivity. Myothermal economy was markedly impaired only in some rats (3/10 in each group), in which the negative inotropic effect of colchicine appeared to be more particularly pronounced. The results indicate that the administration of high doses of colchicine induced intrinsic cardiotoxic effects. Due to its amplitude, such cardiotoxic action may participate in the fatal outcome of acute colchicine poisoning.

Altered Expression of Epidermal Growth Factor Receptor Gene in a Classical Multidrug‐resistant Variant of a Human Cancer Cell Line, KB

A variant clone resistant to high doses of colchicine (KB‐C1) derived from human cancer KB cell line is resistant to various anticancer agents. The KB‐C1 cells were much more resistant to epidermal growth factor and a chimeric toxin, EGF‐Pseudomottas exotoxin (PE), than the parental KB cells. KB‐C1 cells have decreased numbers of EGF‐receptors, though the affinity of the receptors is similar to that in the parental KB cells. A drug‐sensitive revertant (C1‐R2) partially recovered its EGF‐receptor activity. Northern blot analysis showed a decreased level of EGF‐receptor mRNA in KB‐C1 cells, while the multidrug‐resistance gene, mdr‐1, was expressed at very high levels in KB‐C1 cells, but not in KB or C1‐R2 cells. The drug‐resistant cells were less tumorigenic than the parental cells when injected into nude mice. A decreased expression of EGF‐receptor in these cells may be one of the pleiotropic properties of multidrug‐resistant cells and may perhaps represent the basis for their reduced tumorigenicity.

Autoradiographic, ultrastructural and biosynthetic study of the effect of colchicine on enamel matrix secretion and enamel mineralization in hamster tooth germs in vitro

First upper molar tooth germs of two to three days old hamsters were exposed in vitro to colchicine in concentrations ranging between 10 −7 and 10 −4 M in the presence of 45Ca and/or [ 3H]-proline for various times up to 18 h. Enamel mineralization was determined by chemical extraction of in vitro incorporated 45Ca and verified ultrastructurally. Quantitative autoradiography compared with water extracts from total explants radiolabelled with [ 3H]-proline showed a dose-dependent decrease of grain counts over the extracellular enamel to the similar extent as the decrease in radiolabelled amelogenins in water-extracts. It was concluded that water-extracts from total expiants represent amelogenins from the extracellular compartment. Enamel matrix secreted in vitro during exposure to high doses of colchicine failed to mineralize and the complete loss was provoked of the distal parts of the secretory ameloblasts including the distal junctional complexes. Nevertheless, the mineralizing pre-exposure enamel neither hypermineralized nor increased uptake of 45Ca. These data do not support the hypothesis that secretory ameloblasts restrict transepithelial calcium transport by directing most of the calcium ions away from the mineralization front. The biosynthetic data furthermore suggest that enamel matrix proteins, only extractable with guanidine-HCl-EDTA, change their physico-chemical nature during secretory amelogenesis in vitro either during secretion or upon their extracellular mineralization.

Immunocytochemical localization of corticotropin releasing factor (CRF)-like immunoreactivity in the thalamus of the rat

The distribution of corticotropin releasing factor (CRF)-immunoreactive structures in the rat thalamus was studied after treatment with high doses of colchicine (100 μg/100 g b.wt.) with peroxidase-antiperoxidase (PAP) immunocytochemistry in vibratome sections. CRF-immunopositive perikarya were found in the ‘posteromedial complex’ of the thalamus, including the ventromedial, paracentral, mediodorsal, rhomboid, parafascicular nuclei, centrum medianum and ventromedial portion of the posterolateral nucleus. In addition, CRF-containing perikarya were observed in the pretectal and subthalamic nuclei. CRF-immunoreactive processes were seen in most of the medial nuclei of the thalamus. The presence of CRF-immunopositive structures in the thalamus suggests that CRF not only functions as a hypophysiotropic hormone regulating the release of ACTH and β-endorphin from the pituitary, but also as a neurotransmitter or neuromodulator, playing an important role in nociception and analgesia.

Enkephalin systems in diencephalon and brainstem of the rat.

The immunocytochemical distribution of [Leu]enkephalin and an adrenal enkephalin precursor fragment (BAM-22P) immunoreactivity was investigated in the diencephalon and brainstem of rats pretreated with relatively high doses of colchicine (300-400 micrograms/10 microliters intracerebroventricularly). The higher ranges of colchicine pretreatment allowed the visualization of extensive enkephalin-containing systems in these brain regions, some of which are reported for the first time. Immunoreactive perikarya were found in many hypothalamic and thalamic nuclei, interpeduncular nucleus, substantia nigra, the colliculi, periaqueductal gray, parabrachial nuclei, trigeminal motor and spinal nuclei, nucleus raphe magnus and other raphe nuclei, nucleus reticularis paragigantocellularis, vestibular nuclei, several noradrenergic cell groups, nucleus tractus solitarius, as well as in the spinal cord dorsal horn. In addition to the above regions, immunoreactive fibers were also noted in the habenular nuclei, trigeminal sensory nuclei, locus coeruleus, motor facial nucleus, cochlear nuclei, dorsal motor nucleus of the vagus, and hypoglossal nucleus. When adjacent sections of those stained for [Leu]enkephalin were processed for BAM-22P immunoreactivity, it was found that these two immunoreactivities were distributed identically at almost all anatomical locations. BAM-22P immunoreactivity was generally less pronounced and ws preferentially localized to neuronal perikarya. The results of the present as well as the preceding studies (Khachaturian et al., '83) strongly suggest substantial structural similarity between the adrenal proenkephalin precursor and that which occurs in the brain. Also discussed are some differences and parallels between the distribution of [Leu]enkephalin and dynorphin immunoreactivities.

Telencephalic enkephalinergic systems in the rat brain

The comparative distribution of Leu-enkephalin and an enkephalin precursor fragment, BAM-22P, was studied in the rat telencephalon using immunocytochemical procedures. Although the enkephalins have previously been localized to telencephalic neurons, the use of high doses of colchicine in the present study has permitted the visualization of widespread enkephalin-containing systems in cortical-limbic regions. Rats were treated intraventricularly with colchicine (300 to 400 micrograms) to enhance the visualization of neuronal perikarya. Leu-enkephalin immunoreactivity was noted in perikarya as well as processes of neurons in many telencephalic regions including the olfactory bulb, parolfactory areas, olfactory tubercle, basal ganglia, nucleus accumbens, septal nuclei, amygdala, hippocampal formation, and several paleocortical regions such as the piriform, entorhinal, and cingulate cortex, as well as regions of the frontal, parietal, and occipital neocortex. In the majority of regions investigated, analysis of sections adjacent to those processed for Leu-enkephalin histochemistry showed BAM-22P immunoreactivity to be localized to perikarya and processes in the same region. These results suggest that brain enkephalin-positive neurons may possess mechanisms for enkephalin biosynthesis similar to those found in adrenal medullary enkephalin cells.

Limb regeneration blastema cells in the adult newt, Notophthalmus viridescens, give aberrant response to colchicine

This study was designed to investigate the effect of colchicine on the mitotic index (number of mitotic figures per 1000 cells) in the newt limb regeneration blastema. Colchicine doses from 0.1 to 1000 μg per gram body weight and treatment durations from 2 to 72 h were used to determine the dose of colchicine and treatment period resulting in optimum metaphase arrest. A preliminary experiment demonstrated that there was no circadian rhythm of mitosis in the newt limb regeneration blastema. A dose of 5 μg colchicine per gram body weight was found to give the maximum mitotic index; both higher and lower doses had less effect. Very high doses of colchicine resulted in a mitotic index the same as that of the controls. The mitotic index increased with increasing treatment time up to 36 h, after which there was no further increase. In other newt tissues such as duodenal mucosa and liver, a more typical dose–response relationship was observed in that increased colchicine dose led to increased mitotic indices up to the level of lethal doses of colchicine.

Immunocytochemical studies with antisera against Leu-enkephalin and an enkephalin-precursor fragment (BAM-22P) in the rat brain

Rats given high doses of colchicine (300–400 ug, i.c.v.) were used to investigate the comparative distribution of Leu-enkephalin and bovine adrenal medullary enkephalin-precursor fragment (BAM-22P) in the brain. Leu-enkephalin immunoreactive neurons have a widespread distribution throughout the neuraxis. In most areas adjacent section analysis showed BAM-22P and Leu-enkephalin immunoreactive perikarya to be similarly localized. Brain enkephalin biosynthetic mechanisms might therefore be similar to those regulating adrenal medullary enkephalin biosynthesis.