Hypoglycaemic activity of Bixa orellana extract in the dog.

In West Indian folklore, a variety of plants are used for medicinal purposes. One such plant, Bixa orellana (annatto), is commonly used in the treatment of diabetes mellitus. Preliminary studies have shown that a crude annatto seed extract exhibited either glucose lowering or hyperglycaemia-inducing activity depending on how it was further manipulated. This present investigation sought to determine the effects of the glucose-lowering extract on C-peptide and streptozotocin-induced diabetic dogs. This annatto extract was found to decrease blood glucose levels in fasting normoglycaemic and streptozotocin-induced diabetic dogs. In addition, in normal dogs, it suppressed the postprandial rise in blood glucose after an oral glucose load. Interestingly, the extract also caused an increase in insulin-to-glucose ratio in normal dogs. Increased insulin levels were not due to increased insulin synthesis as after 1-h residence time and half-hour postprandial, decreased C-peptide levels was observed. It was concluded that Bixa orellana (annatto) lowered blood glucose by stimulating peripheral utilization of glucose, and it is possible that this glucose-lowering extract might be of pharmacological importance.

The effect of annatto on insulin binding properties in the dog.

Various plants are used in Caribbean folklore for the treatment of a variety of illnesses including diabetes mellitus. Preliminary investigations of several crude plant extracts have indicated that the annatto (Bixa orellana), among others, does in fact exhibit hypoglycaemic properties. This present investigation sought to isolate the hypoglycaemic principle(s) from the crude extract and to determine the mechanism of action. Purification experiments employing thin layer chromatography (TLC) and high performance liquid chromatography (HPLC) resulted in an oil-soluble, partially purified annatto extract. The latter caused a decrease in blood glucose level of 5.62+/-0.13 (n=34) mmol/dL versus 6.31+/-0.12 (n=34) for the control (p<0.01) 1 h after administration. This hypoglycaemia persisted for an additional hour when the oral glucose tolerance test (OGTT) was performed on dogs treated with annatto and compared with the control. Plasma insulin levels measured at 1.0 h showed that there was an increase in plasma insulin levels of 59.57+/-8.3 microIU/mL for the annatto treated dogs versus 40.95+/-5.46 microIU/mL for the control (p<0.05), this elevation persisted throughout the duration of the OGTT which followed. Insulin receptor studies, using a modification of the method of Gambhir et al. done on mononuclear leucocytes and erythrocytes obtained from blood taken 1 h after administration showed that there was an increase in the percentage receptor binding when compared with the control. Insulin affinity results showed that there was an increase of 1.8+/-0.2x10(8) m-1 (n=12) in mononuclear leucocytes for the annatto treated dogs versus 1.2+/-0.2x10(8) m-1 for the control (p<0.05). In the enythrocytes, there was also an increase in affinity from 1.2+/-0.2x10(8) m-1 to 2.3+/-0.2x10(8) m-1 for the control and treated animals, respectively. In conclusion, it can be stated that annatto is responsible for the hypoglycaemic episodes seen in the dogs which was mediated by an increase in plasma insulin concentration as well as an increase in insulin binding on the insulin receptor due to elevated affinity of the ligand for the receptor.

Biphasic alterations of cAMP levels and inhibition of norepinephrine release in iris-ciliary body by bremazocine.

Kappa-opioid receptor agonists have been shown to reduce intraocular pressure in rabbits and monkeys. This study was designed to investigate mechanisms in the iris-ciliary body (ICB) that may be involved in bremazocine (BRE)-induced ocular hypotension in New Zealand White rabbits. Using ICBs, BRE and norbinaltorphimine (nor-BNI), relatively selective kappa-opioid receptor agonist and antagonist, respectively, along with pertussis toxin (PTX), were used to evaluate the effect of 1) kappa-opioid receptors on [(3)H]norepinephrine (NE) release from postganglionic sympathetic neurons, and 2) cAMP accumulation. BRE caused dose-related (0.1, 1, and 10 microM) inhibition of electrically stimulated [(3)H]NE release from ICBs to 77, 57, and 36% of the control, respectively. Nor-BNI antagonized the inhibition of [(3)H]NE release by BRE, while PTX pretreatment limited the suppressive effect of BRE (1 and 10 microM). When used alone, BRE (0.01, 0.1, 1, and 10 microM) caused stimulation of cAMP levels in ICBs, however, similar concentrations caused inhibition of isoproterenol (ISO)-stimulated cAMP production. Pretreatment of ICBs with nor-BNI (10 microM) or PTX (150 ng/ml) antagonized BRE-induced suppression of ISO-stimulated cAMP. These data demonstrate that BRE acts at multiple [prejunctional (neuronal) and postjunctional] sites in the ICB. BRE had a biphasic effect on ISO-stimulated adenylyl cyclase activity; enhancing cAMP levels at low concentrations and inhibiting cAMP production at high concentrations. Based on the modifications induced by PTX pretreatment, the kappa-opioid receptors involved in some of the ocular actions of BRE are linked to a G(i/o) protein.

Elevation of atrial natriuretic peptide levels in aqueous humor of the rabbit by kappa opioid receptor agonists.

Following the topical administration of three kappa agonists (bremazocine, spiradoline and ICI 204448) to rabbit eyes, aqueous humor samples were analyzed for levels of atrial natriuretic peptide (ANP). Bremazocine (BRE) and spiradoline (SPR) elevated aqueous ANP levels in a dose-dependent manner. In contrast, ICI had no significant effect on ANP levels in aqueous humor. Nor-binaltorphimine (nor-BNI), a selective kappa opioid receptor antagonist, was used to assess kappa opioid receptor involvement and glibenclamide, an ATP-sensitive K+ channel blocker, was used to test for the role of ATP-sensitive potassium channels in ANP release. Pretreatment with nor-BNI antagonized the increases in ANP levels observed with both BRE and SPR. Likewise, glibenclamide suppressed the stimulation of ANP secretion by bremazocine. In summary, BRE and SPR increased ANP levels in aqueous humor of rabbits, in part, via activation of K+(ATP) channels that are assumed to be associated with kappa opioid receptors.

Modulation of ocular hydrodynamics and iris function by bremazocine, a kappa opioid receptor agonist.

This study was designed to determine the activity of bremazocine (BRE), a relatively selective kappa opioid receptor agonist, on intraocular pressure (IOP), aqueous humor formation and pupil diameter (PD) in conscious, normal, dark-adapted New Zealand white (NZW) rabbits. IOP was measured in normal and unilaterally sympathectomized rabbits using a calibrated pneumatonometer and the aqueous flow rate was determined by the use of a Fluorotron Master. A masked-design study was conducted in which the rabbits' eyes were treated with BRE topically and unilaterally; the fellow eyes received vehicle. IOP and PD measurements were taken at 0.5 hr and 0 time before BRE and 0.5, 1, 2, 3, 4 and 5 hr post-treatment. Fluorophotometry recordings were taken at 1 hr before and 0.5, 1.5, 2.5 and 3.5 hr after topical application of the drug or vehicle. The effect of the relatively selective kappa opioid receptor antagonist, nor-binaltorphimine (nor-BNI), on bremazocine-induced changes in IOP, PD and aqueous flow was also determined. BRE (10 and 100 micrograms 25 microliters-1 vehicle) produced dose-related, bilateral reductions in IOP, PD and aqueous humor flow. A large increase in IOP (14 mmHg) was observed when BRE (100 micrograms) was applied to sympathectomized eyes. This ocular hypertensive effect was antagonized when the sympathectomized eyes were pretreated with naloxone (200 micrograms), a non-selective opioid receptor antagonist. BRE (10 and 100 micrograms) decreased the aqueous humor flow rate bilaterally by approximately 48 and 60%, respectively, at 0.5 hr after administration to the ipsilateral eye. Nor-BNI (100 micrograms) antagonized the effect of BRE (10 micrograms) on IOP and aqueous flow rates more effectively than on PD. These data indicate that bremazocine causes reductions in IOP by suppressing aqueous flow, but the ocular hypotensive effects are dependent on the presence of intact sympathetic nerves. Antagonism of BRE's effects on aqueous humor dynamics by nor-BNI suggests that the mechanism of IOP and aqueous flow reduction may involve, in part, an action on kappa receptors. Further experiments are necessary to fully define the opioid receptor populations in the ciliary body.

Implementation of pencil kernel and depth penetration algorithms for treatment planning of proton beams.

The implementation of two algorithms for calculating dose distributions for radiation therapy treatment planning of intermediate energy proton beams is described. A pencil kernel algorithm and a depth penetration algorithm have been incorporated into a commercial three dimensional treatment planning system (Helax-TMS, Helax AB, Sweden) to allow conformal planning techniques using irregularly shaped fields, proton range modulation, range modification and dose calculation for non-coplanar beams. The pencil kernel algorithm is developed from the Fermi Eyges formalism and Molière multiple-scattering theory with range straggling corrections applied. The depth penetration algorithm is based on the energy loss in the continuous slowing down approximation with simple correction factors applied to the beam penumbra region and has been implemented for fast, interactive treatment planning. Modelling of the effects of air gaps and range modifying device thickness and position are implicit to both algorithms. Measured and calculated dose values are compared for a therapeutic proton beam in both homogeneous and heterogeneous phantoms of varying complexity. Both algorithms model the beam penumbra as a function of depth in a homogeneous phantom with acceptable accuracy. Results show that the pencil kernel algorithm is required for modelling the dose perturbation effects from scattering in heterogeneous media.

Dose calculation in brachytherapy for a 192Ir source using a primary and scatter dose separation technique.

A dose calculation algorithm for brachytherapy is presented that reduces errors in absolute dose calculation and facilitates new techniques for modelling heterogeneity effects from tissues, internal shields and superficially positioned sources. The algorithm is based on Monte Carlo simulations for specific source and applicator combinations. The dose is scored separately, in absolute units, for the primary and different categories of scatter according to the photon scatter generation. Radial dose distributions for the primary dose and the total scatter dose are parametrized using functions based on simple one-dimensional transport theory. The fitted radial parameters are functions of the angle to the long axis of the source to account for the anisotropy of the dose distribution. The kerma in air at the reference point 1 m from the source is also simulated using Monte Carlo techniques and both the dose and kerma are normalized per source emitted radiant energy. The calculated kerma per radiant energy is used together with the measured reference air kerma rate and the ratio of the dose to the kerma to calibrate the calculated absolute dose rate. Data are presented for an 192Ir cylindrical source, in combination with water, nylon and stainless steel applicators. Values of the radial dose profiles, specific dose rate constants and corrections to the air kerma for attenuation and scatter in air are calculated. Anisotropy functions for the 192Ir source and a water-equivalent applicator are compared to published values. The effects of the applicator wall material on the radial dose distribution are also discussed.

Dose calculations in proton beams: range straggling corrections and energy scaling.

Three-dimensional dose planning systems employing accurate proton transport algorithms are essential for calculating absorbed dose distributions in proton therapy. In this paper, a pencil beam algorithm for the transport of protons in materials of interest for radiation therapy is developed. The Fermi-Eyges multiple-scattering theory is used to derive transport equations for calculating proton fluence and absorbed dose distributions. The multiple-scattering theory of Molière is used to predict mean square scattering angles and to develop an expression for calculating the root mean square (RMS) radial spread of a proton pencil beam, as a function of depth, in an arbitrary scattering material. A correction factor is suggested to account for the decrease in the radial spread at the end of the range due to range straggling. The effects of neglecting large-angle scattering events and the possibility of incorporating such events into the pencil beam algorithm are discussed. An energy scaling technique for determining the water-equivalent surface energy at a given depth in a heterogeneous scattering medium is developed. The water-equivalent energy, giving the same Molière scattering parameter B in water, is determined and the 1/e angle in water is scaled to the appropriate width in the scattering material. By using stored analytically or Monte Carlo calculated pencil beam distributions in water, the large-angle single-scattering events may be incorporated by approximating the scattering in an arbitrary material by the scattering in water for protons of the appropriate water-equivalent surface energy.

Cell type dependent effectiveness of tumour cell inactivation by radiation with increased ionisation density.

Radiation with increased ionization density, LET, will in the near future be applied in targeted radiotherapy and has already, to some degree, been applied in external radiotherapy with accelerated ions. There are indications from the literature that different types of cells are sensitized differently when the LET is increased. Radiation with three different ionisation densities was applied in the present study; 0.8 keV/microns photons from a reference 60Co source, 6 keV/microns helium ions from the entrance region of a monoenergetic helium ion beam and 25 keV/microns from a range modulated helium beam. Three types of cultured cells were analysed: human colon carcinoma LS-174T, human glioma U-343MG and hamster embryonic lung V79-379A. There were interesting differences between the cell types; the LS-174T cells showed strong sensitization already at an LET of 6 keV/microns and there was nearly no difference between the shape of the survival curves after irradiation with 6 or 25 keV/microns. The V79-379A cells were less sensitive to the 6 keV/microns radiation quality and the survival curve was, in this case, very similar to the reference 60Co survival curve. However, the V79-379A cells were sensitized when exposed to the 25 keV/microns helium ions. The U-343MG cells were intermediate in the sense that the 6 keV/microns curve fell in between the 60Co and the 25 keV/microns curves. These variations indicate that different types of cells react differently to changes in LET and that this is probably of special interest for intermediate LET radiation. Some cells might be easily sensitized by intermediate LET qualities while others might be more resistant and this is of importance for the future optimization of radiation treatment with both targeted agents and accelerated ions.

Patient positioning for fractionated precision radiation treatment of targets in the head using fiducial markers.

When irradiating targets in the brain, an accurately localised dose is often needed. One crucial moment to achieve this is the positioning of the patient. We have developed a positioning method where the patient is immobilised with a bite block and a head mould, and where the position of the target is determined by X-ray imaging of fiducial markers that are placed in the patient's skull. A method for computing the transformation needed to move the target from the observed to the prescribed position and orientation is described. This method uses the information from two orthogonal X-ray images and takes measurement errors and data from three or more markers into account. Results from using the method clinically in proton beam therapy are given.

Relative biological effectiveness of intermediate energy protons. Comparisons with 60Co gamma-radiation using two cell lines.

Range modulated proton beams are used for radiotherapy of malignant tumours at several accelerator laboratories with the aim of introducing proton therapy as a clinical hospital-based therapy modality. Due to the finite range and the sharpness of the dose gradients, the dose to well defined target volumes can be raised without excessive irradiation of non-target tissue. The prescribed proton doses are determined in part on the basis of the relative biological effectiveness (RBE) of the particular radiation quality. In this study, RBE values were determined for a proton beam with a maximal range of 33 mm, which corresponds to an energy of approximately 67 MeV. The range modulated depth-dose distribution, with a 20 mm extended Bragg peak, was mainly designed for high precision treatment of small targets such as uveal melanomas. The tested cell lines, LS-174T and V79-379A, were chosen because of their suitability for clonogenic assays. The cells were irradiated with single doses in the range 2-10 Gy at different depths in the extended peak region of the range modulated proton beam. RBE values were determined by comparing the doses needed to obtain the same reduction in colony formation (0.5, 0.1 and 0.01) as with the reference 60Co gamma source. The mean RBE value was 1.22 with a standard deviation of 0.08. The variations depended on both cell type and on the survival levels considered.(ABSTRACT TRUNCATED AT 250 WORDS)