Carrying and spine loading.

The advantages and disadvantages of different methods of carrying objects on spine loading are still not fully understood. Previous studies have either examined the effects of carrying using physiological measures or examined isolated spine segments using biomechanical models. Additionally, most studies have been restricted to only a small number of carrying conditions. Very few studies have attempted to examine the various factors influencing spine loading together. To improve understanding of interacting factors on carrying, this study assessed the lumbar spine loads of 16 subjects as they assumed six styles of carrying at two weight levels and two activity levels (walking vs. standing). Concurrent with each trial, a subject-specific biomechanical model was used to assess spine forces over the full lumbar spine. Most carrying methods in the trials resulted in relatively low levels of spine loading. Anterior/posterior (A/P) shear loading was the only spine-loading dimension that reached biomechanically meaningful levels. Two carrying conditions, with bins carried in front of the body, significantly increased A/P shear compared with other carrying styles. This increase appeared to be due to the greater moment arms occurring in these conditions. Many of the other carrying styles produced A/P shears that were similar to those observed when carrying nothing at all. Of all the tasks, the backpack carry characteristically produced especially low spine loads. The findings of the study suggest that to achieve optimal carrying in terms of spine loading, loads should be positioned close to the body, even when carrying relatively light loads.

Pheromones enhance somatosensory processing in newt brains through a vasotocin-dependent mechanism.

We tested whether the sex pheromones that stimulate courtship clasping in male roughskin newts do so, at least in part, by amplifying the somatosensory signals that directly trigger the motor pattern associated with clasping and, if so, whether that amplification is dependent on endogenous vasotocin (VT). Female olfactory stimuli increased the number of action potentials recorded in the medulla of males in response to tactile stimulation of the cloaca, which triggers the clasp motor reflex, as well as to tactile stimulation of the snout and hindlimb. That enhancement was blocked by exposing the medulla to a V1a receptor antagonist before pheromone exposure. However, the antagonist did not affect medullary responses to tactile stimuli in the absence of pheromone exposure, suggesting that pheromones amplify somatosensory signals by inducing endogenous VT release. The ability of VT to couple sensory systems together in response to social stimulation could allow this peptide to induce variable behavioural outcomes, depending on the immediate context of the social interaction and thus on the nature of the associated stimuli that are amplified. If widespread in vertebrates, this mechanism could account for some of the behavioural variability associated with this and related peptides both within and across species.

Impact on renal function after endovascular aneurysm repair with uncovered supra-renal fixation assessed by serum cystatin C.

OBJECTIVE: Supra-renal fixation in endovascular aneurysm repair (SR-EVR) is used to improve the proximal seal of aortic stent grafts and appears to have minimal effect on serum creatinine. Serum cystatin C (CC) is a more sensitive marker of renal injury and, unlike creatinine, is unaffected by non-renal influence. The aim of this study was to assess the true renal effect of SR-EVR using this superior renal index. METHODS: Consecutive patients undergoing SR-EVR were prospectively recruited and compared to control groups undergoing open aneurysm repair (OR) and colorectal resection (CR). Serum CC and creatinine clearance (CrC) were determined pre-operatively and at 3, 6 and 12 months post-surgery. Renal function was compared using analyses of covariance (ANCOVA). RESULTS: Sixty-five patients (M:F; 52:13, median age 74 years) were enrolled (24 SR-EVR, 28 OR, 13 CR). Pre-operative renal function and risk factors were comparable (CC 1.04mg/l, SR-EVR; 0.96mg/l, OR; 0.97mg/l, CR). Adjusting for baseline renal function, there was no significant difference in CC or CrC between study and both control groups at 3, 6 or 12-months post-operatively. CONCLUSION: Using cystatin C as a more sensitive renal index, there was no detectable evidence of kidney dysfunction at up to one-year following EVR with uncovered bare-metal supra-renal fixation.

Bis(tBuSATE) phosphotriester prodrugs of 8-azaguanosine and 6-methylpurine riboside; bis(pom) phosphotriester prodrugs of 2'-deoxy-4'-thioadenosine and its corresponding 9alpha anomer.

As an extension of previous work with bis(POM) nucleotide prodrugs, we report the synthesis and biological evaluation in tumor cell culture of the bis(pivaloyloxymethyl) phosphotriester prodrug of slightly cytotoxic 2'-deoxy-4'-thioadenosine and its alpha-anomer. We have experienced need for an alternative phosphate masking group, particularly with purine nucleosides. Accordingly, we report synthesis and biological evaluation of the bis(tBuSA TE) phosphotriester prodrugs of 8-azaguanosine and 6-methylpurine riboside, nucleoside analogs with moderate to significant cytotoxicity. All four prodrugs were examined in tumor cell culture in parallel with the parent nucleosides. Synthetic routes and biological data are presented.

Forebrain influences on brainstem and spinal mechanisms of copulatory behavior: a current perspective on Frank Beach's contribution.

In a 1967 Physiological Reviews paper, Frank Beach put forth four propositions regarding forebrain and hormonal control of brainstem-spinal mechanisms of copulatory behavior. Simply stated, he proposed that: 1) the forebrain exerted an inhibitory control over species-typical copulatory reflexes through descending effects on brainstem-spinal mechanisms and 2) gonadal hormones influence these reflexes largely by actions on forebrain control processes rather than by direct effects on the brainstem or spinal cord. This theoretical scheme was of great heuristic significance during the subsequent two decades of research, which has largely supported and delineated in greater mechanistic detail the processes Beach hypothesized to exist. This subsequent research has also shown the central nervous system actions of gonadal hormones to be more widespread and complex than Beach proposed. Some of these recent research findings are presented, with emphasis on neurophysiological studies which have identified hormone-induced functional changes in forebrain and brainstem neurons. It is proposed that these functional changes may represent a mechanism for the behavior-controlling actions of hormones that were hypothesized by Beach.

Preparation and purification of L-(+/-)-5-formyl-5,6,7,8-tetrahydrofolic acid.

Reinvestigation of the conversion of folic acid to leucovorin [L-(+/-)-5-CHO-THF] led to improved methods for the synthesis of this drug, which is suitable for clinical use. Also, methods were developed for the chromatographic and nonchromatographic purification of less pure samples of L-(+/-)-5-CHO-THF.

Synthesis of potential anticancer agents: imidazo[4,5-c]pyridines and imidazo[4,5-b]pyridines.

The 1,2-dihydropyrido[3,4-b]pyrazines (1) are mitotic inhibitors with significant antitumor activity in mice. Also, the active imidazo[4,5-b]pyridine 3 was shown to cause the accumulation of cells at mitosis. Routes were developed for the synthesis of congeners of 3 by cyclization of 4-(substituted amino)-5,6-diaminopyridines with ethyl orthoformate. Oxidative cyclization of either 4,5- or 5,6-diaminopyridines with aryl aldehydes produced the [4,5-c] and [4,5-b] imidazopyridine ring systems, respectively. The latter reaction with 6-(substituted amino)-4,5-diaminopyridines gave imidazo[4,5-c]pyridine ring analogues of 1. Biological studies indicated that the target compounds were less active than 1 and 3.

New synthesis of N-[4-[[(2-amino-4(3H)-oxopyrido[3,2-d]pyrimidin-6-yl)methyl]amino]benzoyl]-L-glutamic acid (8-deazafolic acid) and the preparation of some 5,6,7,8-tetrahydro derivatives.

Previously, 8-deazafolic acid (17) was shown to be a potent inhibitor of the folate-dependent bacteria, Streptococcus faecium (ATCC 8043) and Lactobacillus casei (ATCC 7469), and to have activity against lymphoid leukemia L1210 in mice. To examine the 5,6,7,8-tetrahydro derivatives, a new synthesis of 17 was developed from 8-deaza-2,4-dichloro-6-methylpteridine. Treatment of the latter with aqueous base gave the corresponding pteridin-4(3H)-one, which was aminated with ammonia to give 8-deaza-6-methylpterin (9). Bromination of 9 gave mainly 8-deaza-6-(tribromomethyl)pterin, which on reaction with p-aminobenzoyl-L-glutamic acid resulted in the formation of the 9-oxo derivative of 17. In contrast, bromination of the 2-acetyl derivative of 9 gave mainly the corresponding 6-(bromomethyl)pterin, which was converted to 17 in 23% yield (from 9). Hydrogenation of 17 at atmospheric pressure and room temperature was unsuccessful either in a basic medium or formic acid. In trifluoroacetic acid, overreduction occurred to give a mixture containing 8-deaza-5,6,7,8-tetrahydro-6-methylpterin and the 5,6,7,8-tetrahydro derivative of 17. The latter was characterized by conversion to the methenyl analogue 21, which was also prepared by hydrogenation of the 10-formyl derivative of 17. Treatment of 21 with hydroxide gave 8-deaza-10-formyl-5,6,7,8-tetrahydrofolic acid. Compound 21 showed cytotoxicity to cultured H.Ep.-2 cells and was tested as an inhibitor of bovine dihydrofolic reductase. Lineweaver-Burk analysis indicated inhibition competitive with dihydrofolate.

New anticancer agents: synthesis of 1,2-dihydropyrido[3,4-b]pyrazines (1-deaza-7,8-dihydropteridines).

Reaction of alpha-aminoacetophenone oximes (2) with ethyl 6-amino-4-chloro-5-nitropyridine-2-carbamate (1) gave ethyl 6-amino-5-nitro-4-[(2-oxo-2-phenylethyl)amino]pyridine-2-carbamate oximes (3), which were hydrolyzed under acidic conditions to give the corresponding ketones (4). Related pyridines substituted with a keto side chain were prepared from 1 and 1,3-diaminopropanone oximes and by oxidation of the side-chain hydroxy group of ethyl 6-amino-4- [[3-(N-methyl-N-phenylamino)-2-hydroxypropyl]amino]-5-nitropyridine-7- carbamates (6). Catalytic hydrogenation of the nitro group of 4 over Raney nickel in a large volume of ethanol gave the 1-deaza-7,8-dihydropteridines (7). Several of the oximes 3 were successfully hydrogenated to give 7 directly. The resulting 1-deaza-7,8-dihydropteridines showed potent cytotoxicity against cultured L1210 cells and significant anticancer activity against lymphocytic leukemia P-388 in mice. These biological activities are attributed to the accumulation of cells at mitosis.

Synthesis of pseudo cofactor analogues as potential inhibitors of the folate enzymes.

Reaction of 5,6,7,8-tetrahydrofolic acid (THF,7) with phosgene, thiophosgene, and cyanogen bromide gave the bridged derivatives, 5,10-(CO)-THF (8), 5,10-(CS)-THF (9), and 5,10-(C = NH)-THF (11), respectively. Catalytic hydrogenation of 10-(chloroacetyl)folic acid (2) gave 5,10-(CH2CO)-THF (12). A similar reaction with 10-(3-chloropropionyl)folic acid (3) gave 10-(ClCH2CH2CO)-THF (14) rather than 5,10-(CH2CH2CO)-THF (13). In the catalytic hydrogenation of 10-ethoxalylfolic acid (5), the initial product 10-(EtO2CCO)-THF (22) rearranged readily to give 5-(EtO2CCO)-THF (21). Acylation of THF with chloroacetyl chloride gave a N5,N10-diacylated product (18 or 19), which could not be converted to 5,10-COCH2)-THF (17). Reductive alkylation of THF with glyoxylic acid and 5-hydroxypentanal, respectively, gave 5-(HO2CCH2)-THF (24) and 5-[HO(CH2)5]-THF (25). Reductive dialkylation of THF with formaldehyde gave 5,10-(CH3)2-THF (27), whereas glyoxal gave 5,10-CH2CH2)-THF (10). Also, both folic acid and 5-(CHO)-THF were reductively alkylated with formaldehyde to give 10-methylfolic acid (6) and 5-(CHO)-10-(CH3)-THF (28), respectively. These compounds were tested as inhibitors of the enzymes involved in folate metabolism and for activity against lymphocytic leukemia P388 in mice.

1,2-Dihydropyrido[3,4-b]pyrazines: structure-activity relationships.

Certain derivatives containing the 1,2-dihydropyrido[3,4-b]pyrazine (1-deaza-7,8-dihydropteridine) ring system are active against experimental neoplasms in mice. The mechanism of action of these agents has been attributed to the accumulation of cells at mitosis. Identification of the structural features that are necessary for activity was accomplished by evaluation of modified 1-deazapteridines and ring and ring-opened analogues. Relative to ethyl 4-amino-1-deaza-7,8-dihydro-6-[(N-methylanilino)methyl]pteridine-2-carbamate (11) and the corresponding 6-phenyl compound (12), no antitumor activity was observed with 7,8-dihydropteridines, 3-deaza-7,8-dihydropteridines, and the corresponding heteroaromatic compounds. Also, activity was diminished or destroyed when 1-deaza-7,8-dihydropteridines were oxidized to 1-deazapteridines or reduced to 1-deaza-5,6,7,8-tetrahydropteridines. In addition, replacement of the 4-amino group with other substituents destroyed activity. The presence of a 6-substituent containing an aryl group appeared to be necessary for activity, which was increased when a methyl group was substituted at the 7-position.

Structure-based design of inhibitors of purine nucleoside phosphorylase. 1. 9-(arylmethyl) derivatives of 9-deazaguanine.

Purine nucleoside phosphorylase (PNP, EC 2.4.2.1) is a salvage enzyme important to the T-cell-mediated part of the immune system and as such is an important therapeutic target. This paper describes the design, synthesis, and enzymatic evaluation of potent, competitive inhibitors of PNP. Potential inhibitors were designed using the three-dimensional structure of the enzyme in an iterative process that involved interactive computer graphics to model the native enzyme and complexes of it with the inhibitors, Monte Carlo-based conformational searching, and energy minimization. Studies of the enzyme/inhibitor complexes were used to determine priorities of the synthetic efforts. The resulting compounds were then evaluated by determination of their IC50 values and by X-ray diffraction analysis using difference Fourier maps. In this manner, we have developed a series of 9-(arylmethyl)-9-deazapurines (2-amino-7-(arylmethyl)-4H-pyrrolo[3,2-d]-pyrimidin-4-ones) that are potent, membrane-permeable inhibitors of the enzyme. The IC50 values of these compounds range from 17 to 270 nM (in 1 mM phosphate), with 9-(3,4-dichlorobenzyl)-9-deazaguanine being the most potent inhibitor. X-ray analysis explained the role of the aryl groups and revealed the rearrangement of hydrogen bonds in the binding of the 9-deazaguanines in the active site of PNP relative to the binding of the 8-aminoguanines that results in more potent inhibition of the enzyme.

Structure-based design of inhibitors of purine nucleoside phosphorylase. 3. 9-Arylmethyl derivatives of 9-deazaguanine substituted on the methylene group.

X-ray crystallography and computer-assisted molecular modeling (CAMM) studies aided in the design of a potent series of mammalian purine nucleoside phosphorylase (PNP) inhibitors. Enhanced potency was achieved by designing substituted 9-(arylmethyl)-9-deazaguanine analogs that interact favorably with all three of the binding subsites of the PNP active site, namely the purine binding site, the hydrophobic pocket, and the phosphate binding site. The most potent PNP inhibitor prepared during our investigation, (S)-9-[1-(3-chlorophenyl)-2-carboxyethyl]-9-deazaguanine (18b), was shown to have an IC50 of 6 nM, whereas the corresponding (R)-isomer was 30-fold less potent.

Structure-based design of inhibitors of purine nucleoside phosphorylase. 2. 9-Alicyclic and 9-heteroalicyclic derivatives of 9-deazaguanine.

Alicyclic and heteroalicyclic derivatives of 9-deazaguanine (2-amino-1,5-dihydro-4H-pyrrolo[3,2-d] [pyrimidin-4-one) are, with one exception, potent inhibitors of purine nucleoside phosphorylase (PNP) equaling the corresponding 9-arylmethyl derivatives previously investigated. The mode of binding of these compounds to PNP was determined by X-ray crystallography.

Steroid-neuropeptide interactions that control reproductive behaviors in an amphibian.

Investigations into the neuroendocrine regulation of reproductive behaviors in an amphibian (Taricha granulosa) reveal the same basic repertoire of chemical messengers as regulators of male behaviors in other vertebrates. These studies have identified seasonal neural interactions between gonadal steroids and neuropeptides that facilitate male courtship behavior. In addition, this species has served to elucidate how stress-induced suppression of courtship is mediated by corticosterone action through a neuronal membrane receptor and subsequent, rapid neurophysiological effects. These findings indicate that a principal mechanism by which steroids and neuropeptides control male reproductive behavior is the modulation of neural processing of specific sensory stimuli.

Echophonocardiographic diagnosis of left ventricular pseudoaneurysm.

We report the presence of an unusual systolic murmur associated with a traumatic left ventricular pseudoaneurysm. Echophonocardiographic studies showed the murmur to begin at the first heart sound, but end well before the second heart sound. It seems likely that the murmur is caused by the systolic flow of blood from the left ventricle into the relatively noncompliant pseudoaneurysm. The echocardiographic scan of the left ventricle demonstrated a relatively echo-free space posterior to the left ventricular wall, supporting the diagnosis of pseudoaneurysm, which has confirmed with angiographic studies and at surgery. These findings indicate that a combination of noninvasive techniques is useful in establishing this diagnosis.

Low-sexually performing rams but not male-oriented rams can be discriminated by cell size in the amygdala and preoptic area: a morphometric study.

Brain regions of male sheep behaviorally classified as high-sexually performing (n=10), low-sexually performing (n=8) or male-oriented (n=9) were examined to determine if differences in reproductive behavior were associated with differences in density or sizes of neurons. High-sexually performing rams actively mounted estrous ewes, low-sexually performing rams failed to mount or had long latencies to mounting estrous ewes, and male-oriented rams mounted other rams in preference to ewes in estrus. Cell densities and sizes were quantified in Nissl stained sections through the medial amygdala (meAMY), preoptic area (POA), bed nucleus of the stria terminalis (BNST), ventromedial hypothalamic nucleus (VMH), lateral geniculate nucleus (LG) and medial geniculate nucleus (MG). Multivariate discriminant analysis based on soma sizes within nuclei of known importance for reproductive behavior and/or gonadotropin release (meAMY, POA, BNST and VMH) discriminated (Wilks Lambda P<0.05) low-performing rams from high-performing and male-oriented rams, but did not discriminate (Wilks Lambda P=0.14) between high-performing and male-oriented rams. Cell size in the parvocellular and magnocellular layers of the LG along with cells of the MG, structures without a specific role in reproduction, did not discriminate any of the three behaviorally defined groups of rams (Wilks Lambda P=0.57). Density of cells present in structures important for the display of reproductive behavior (POA, meAMY, BNST) and/or gonadotropin release (POA, VMH) had no discriminating power nor did density of cells in structures important for the processing of visual (LG) or auditory (MG) stimuli. In conclusion, significant differences in sizes of cells located within nuclei that are specifically important for the display of male reproductive behavior were found in low-sexually performing rams compared to high-sexually performing and male-oriented rams. These differences may result from neuron development in utero or occur later as a consequence of endocrine factors or behavioral experience. Neuronal cell size is a critical variable that determines excitability to synaptic inputs because cell surface area varies exponentially with cell diameter. Relatively small differences in neuron diameter could relate to functionally important differences in neuronal excitability.