Timothy Othman

Oligodendrocytes express functional A1 adenosine receptors that stimulate cellular migration.

A1 adenosine receptors (A1ARs) exert important effects in the central nervous system. However, the expression and function of A1ARs in oligodendrocyte precursor cells (OPCs) and oligodendrocytes (OLGs) is unclear. To address this issue, we examined A1AR expression during different stages of oligodendrocyte development. Radioreceptor studies showed that membranes prepared from OPCs and OLGs expressed high‐affinity A1ARs with Kd values of 1.35 ± 0.33 and 1.2 ± 0.27 nM for [3H]CCPA, 1.17 ± 0.24 and 1.4 ± 0.34 nM for [3H]DPCPX, respectively. Bmax values were 64.31 ± 6.14 and 75 ± 6 fmol/mg protein for [3H]CCPA, and 153 ± 12 and 205 ± 17.8 fmol/mg protein for [3H]DPCPX, respectively. Activation of A1ARs using N6‐cyclopentyladenosine (CPA) reduced both forskolin‐ and N‐ethylcarboxyamidoadenosine (NECA)‐stimulated cAMP accumulation, but did not affect basal cAMP levels. Activation of A1ARs by CPA stimulated OPC migration, but did not affect cell viability, proliferation, or differentiation. These results show that OPCs and OLGs express functional A1ARs that can stimulate the migration of OPCs.

A1 adenosine receptor activation induces ventriculomegaly and white matter loss.

A1 adenosine receptors (A1ARs) are widely expressed in the brain during development. To examine whether A1AR activation can alter postnatal brain formation, neonatal rats from postnatal days 3 to 14 were treated with the A1AR agonist N6-cyclopentyladenosine (CPA) in the presence or absence of the peripheral A1AR antagonist 8-(p-sulfophenyl)-theophylline (8SPT). CPA or CPA + 8SPT treatment resulted in reductions in white matter volume, ventriculomegaly, and neuronal loss. Quantitative electron microscopy revealed reductions in total axon volume following A1AR agonist treatment. We also observed reduced expression of myelin basic protein in treated animals. Showing that functional A1ARs were present over the ranges of ages studies, high levels of specific [3H]CCPA binding were observed at PD 4, 7 and 14, and receptor-G protein coupling was present at each age. These observations show that activation of A1ARs with doses of CPA that mimic the effects of high adenosine levels results in damage to the developing brain.

Cytoskeletal protein 4.1G binds to the third intracellular loop of the A1 adenosine receptor and inhibits receptor action.

To identify binding partners of the A1AR (A1 adenosine receptor), yeast two-hybrid screening of a rat embryonic cDNA library was performed. This procedure led to the identification of erythrocyte membrane cytoskeletal protein (represented as 4.1G) as an A1AR-binding partner. Truncation studies revealed that the C-terminal domain of 4.1G was essential for binding to A1ARs and that the C-terminal domain of 4.1G and the third intracellular loop of A1ARs interacted. A1AR-4.1G interaction was also confirmed in studies using brain tissue. Studies in HEK-293 (human embryonic kidney 293) cells and Chinese-hamster ovary cells showed that 4.1G interfered with A1AR signal transduction, as 4.1G reduced A1AR-mediated inhibition of cAMP accumulation and intracellular calcium release. 4.1G also altered cell-surface A1AR expression. These observations identify 4.1G as a novel A1AR-binding partner that can regulate adenosine action.

Cytoskeletal protein 4.1G is a binding partner of the metabotropic glutamate receptor subtype 1α

Recent evidence suggests that cytoskeletal proteins play important roles in the clustering and anchoring of glutamate receptors to the cell surface membrane. To examine further this issue, we tested for direct interactions between the metabotropic glutamate receptor subtype 1α (mGlu1α) and 4.1G, which is a member of the erythrocyte membrane, cytoskeletal protein 4.1 family. First, co‐localization of 4.1G and mGlu1α was observed in cultured hippocampal neurons. Second, in transiently transfected HEK 293 cells and in whole rat brain tissue, direct interactions between mGlu1α and 4.1G were observed. Third, we were able to identify the C‐terminal tail of mGlu1α as an essential region for mGlu1α–4.1G interactions. Fourth, 4.1 G influences mGlu1α‐mediated cAMP accumulation. Finally, we found that 4.1G increases the ligand‐binding ability of mGlu1α and alters its cellular distribution. These observations identify 4.1G as a novel binding partner of mGlu1α that can regulate the action of mGlu1α.

Hyperthermic Enhancement of the Apoptotic and Antiproliferative Activities of Paclitaxel

The antineoplastic agent paclitaxel (PTX), a microtubule-stabilizing agent, is known to arrest cell cycle progression and induce apoptosis. Mild hyperthermia (HT) also disrupts the microtubule system and triggers apoptosis. We therefore investigated whether concurrent exposure of murine breast cancer cells to 10 µmol/l PTX and 43°C HT will promote improved anticancer effects. To do this, we exposed FM3A murine cancer cells to: (1) 10 µmol/l PTX for 1 h at 37°C followed by exposure at 43°C HT for 1 h; (2) 10 µmol/l PTX at 37°C for 2 h; (3) 37°C for 1 h followed by 43°C HT for 1 h, and (4) untreated cells at 37°C for 2 h which served as the control. Treatment No. 1 resulted in an enhanced cell cycle arrest, apoptosis and cytotoxicity. Exposure to 43°C HT alone or 10 µmol/l PTX alone induced lesser apoptosis and cytotoxicity than the two treatments concurrently applied. The apoptotic cell death occurred in a time-dependent manner as follows: (1) concurrently applied 43°C HT and 10 µmol/l PTX (5.6 ± 0.5, 16.5 ± 2 and 27.6 ± 1%); (2) 43°C HT alone (4.3 ± 1, 6.6 ± 0.3 and 12.7 ± 1%) and (3) 10 µmol/l PTX alone (4.4 ± 0.3, 8.6 ± 1 and 12.8 ± 1%) at 1, 6 and 24 h postexposure respectively compared to control of 2.0%. These data indicate that while both HT and PTX can individually induce apoptosis and antiproliferation in FM3A cancer cells, they may offer synergistic benefits when used concurrently.

Relationships Between IgG, IgM, IgE and Resistance to Reinfection During the Early Phase of Infection with Clonorchis sinensis in Rats

An enzyme linked immunosorbent assay (ELISA) was used to study the correlation between the levels of IgG, IgM and IgE immunoglobulin isotypes and resistance to re‐infection in rats during the first month of infection with Clonorchis sinensis. Rats were infected with Clonorchis sinensis (primary infection), and then treated with praziquantel on the 1st, 3rd, 7th, 14th and 28th day post infection (p.i.). To measure resistance, rats were re‐infected with C. sinensis (secondary infection), 2 weeks after the treatment and worms were recovered 4 weeks later. During the primary infection, significantly increased levels of IgG isotype were observed on days 14 and 28 p.i. (P < 0.001) and IgM levels were significantly increased on 3rd and 28th day (P < 0.001). During the secondary infection, significantly increased levels of IgG isotype were found from 3rd to 28th day and IgE isotype on 7th and 14th day (P < 0.01) while significant levels of IgM were found on the 3rd and 28th day (P < 0.05). Furthermore, significant differences of worm numbers between infected and control group was found on the 14th and 28th day (P < 0.001). An inverse correlation between the IgG levels and the resistance to re‐infection was also observed (r = − 0.948, P = 0.004), indicating that the resistance to reinfection is highly associated with the levels of IgG during the early phase of infection, and then with the IgM and IgE.

Anticancer efficacies of doxorubicin, verapamil and quercetin on FM3A cells under hyperthermic temperature

Hyperthermia (HT) in combination with anticancer drugs (ACDs) had proven to more efficacious in various cancers, although efficacies vary according to chemotherapeutic compounds and cancer types. Presently there are few data that compares anticancer efficacies among ACDs under hyperthermic conditions. Therefore, we selected three commonly used ACDs (quercetin, verapamil and doxorubicin) and compared their antitumor effects when each was treated with 43°C HT exposure. Firstly, FM3A, a murine breast cancer cell line, was treated with each ACD for 1 h followed by 43°C exposure for additional 1 h, and examined the effects of: 1) each drug, 2) 43°C HT exposure, and 3) the combination of each drug and 43°C HT exposure for 1, 6 and 24 h. The determined overall effects on FM3A cells were arrested cell proliferation, clonogenic efficiency and apoptosis. Pre-treatment of FM3A cells to each ACD followed by 43°C HT exposure produced greater antitumor effects including suppressed cell proliferation, reduced clonogenic efficiency and increased apoptotic cell death, compared to ACD treatment or HT exposure alone. Apoptotic cell death occurred in a time-dependent manner. Among the ACDs, antitumor efficacies varied in the order of doxorubicin > verapamil > quercetin. It was concluded that heat exposure during ACD treatment of caner cells may be an important factor to get a better antitumor benefit, even though this benefit may differ from one drug to another.

Central and Peripheral Control of Sweating Mechanisms: Modifications by Thermal Acclimatization and Physical Training

Sweating is a heat loss response that is critical for improved physical performance and safety in extremely hot conditions. It is centrally regulated by the preoptic area and anterior hypothalamus and peripherally transmitted by sympathetic sudomotor innervation, with acetylcholine (ACh) as the primary neuroglandular transmitter. Modification of sweating activity through heat exposure or physical training is a physiological tactic for improved tolerance when individuals are challenged with exogenous or endogenous heat. A short-term heat challenge produces a lower resting and slower increase in body temperature as well as enhanced sweating response, while long-term heat exposure results in decreased sweat output. Cold acclimation results in reduced thermoneutral and skin temperatures, lowered cold sensation, and reduced metabolic heat production. Physical training induces higher sweat output by means of greater sweat output per activated sweat gland, a shorter lag phase for sweating, an increased number of activated sweat glands, and a higher rate of skin blood flow.

Anatomical and neurochemical peculiarities of the pika retina: basis for lack of circadian rhythm of core temperature

We have previously reported a complete lack of circadian rhythm in the body temperature of pikas in contrast to other lagomorphs. In this present study, the anatomical and neurochemical findings by immunohistochemical, photo and electron microscopic methods reveal that the photoreceptor system of this animal is poorly developed. This probably explains their stable core body temperature which help them survive in cold temperatures.