Body mass index and occupational accidents among health care workers in a large university hospital.

INTRODUCTION: Obesity is associated with a number of chronic diseases such as cardiovascular diseases and cancers. The association of obesity with occupational accidents has been suggested although the evidence is less convincing. The objective of the study is to analyse the relationship between BMI values and ergonomic accidents in a large University Hospital. METHODS: The relationship between body mass index (BMI) and the incidence of ergonomic occupational accidents over a period of 8 years was investigated in a cohort of employees of a large University Hospital, covering almost 27,000 person-years of observation. This relationship was stratified according to the variables age, gender, functional status within the organization and work schedule (part-time or full time). Height and weight were objectively measured, demographic data were obtained from the human resource department and the registration of ergonomic accidents was carried out by the safety and prevention department of the hospital. RESULTS: The number of ergonomic accidents, expressed as number/1000 person-years was higher for female employees compared to male employees, increased with age and markedly increased from functional class A (leading or expert function and higher educational level) to D (executive function in patient care and technical department). However, the incidence of ergonomic accidents accompanied by loss of working time was not significantly associated with BMI, independently of age and gender. In addition, the type of accident and the severity of the accidents expressed as the number of days absent from work were unrelated to BMI. CONCLUSION: No independent relationship between BMI and the incidence of ergonomic accidents could be identified in our cohort. Tailoring working conditions to individual BMI levels is not recommended.

The drone ambulance [A-UAS]: golden bullet or just a blank?

Defibrillation within the first minutes after sudden cardiac arrest can save many quality-adjusted life years. Yet, despite enormous investments, 'healthcare' is still unable to provide this for the majority of patients. Emergency Medical Services often have a too long mean response time and many issues surround Public Access Defibrillation programs. In this article we argument that AED-equipped drones could be the 'magic bullet'. They are easily deployed and fast, and have a relatively low operational cost. As such they could rapidly bring an AED next to the victim, irrespective of most geographical circumstances, give visual feedback and situational awareness to the EMS dispatcher and thus assist a bystander to provide better CPR. Although there are many real-life barriers to actual deployment, we argument these might all get solved once we have solved the described technological issues.

Desensitization of muscarinic receptors.

When Chinese hamster ovary cells transfected with the gene for M(3)-muscarinic receptors were stimulated with carbachol continuously for 30 min, the response at the end of the stimulation period was about 20% of the early response (2-3 min after the start of the stimulation). Long-term treatment of the cells with phorbol ester abolished the response completely while desensitization was significantly reduced upon pre-treatment of the cells with GF109203X, antisense oligonucleotide against the alpha-isoform of protein kinase C and wortmannin. We conclude that in the Chinese hamster ovary expression system, desensitization of M(3)-muscarinic receptors is dependent on a fast feedback loop including the alpha-isoform of protein kinase C.

Prefoldin recognition motifs in the nonhomologous proteins of the actin and tubulin families.

Nascent actin and tubulin molecules undergo a series of complex interactions with chaperones and are thereby guided to their native conformation. These cytoskeletal proteins have the initial part of the pathway in common: both interact with prefoldin and with the cytosolic chaperonin containing tailless complex polypeptide 1. Little is understood with regard to how these chaperones and, in particular, prefoldin recognize the non-native forms of these target proteins. Using mutagenesis, we provide evidence that beta-actin and alpha-tubulin each have two prefoldin interaction sites. The most amino-terminally located site of both proteins shows striking sequence similarity, although these proteins are nonhomologous. Very similar motifs are present in beta- and gamma-tubulin and in the newly identified prefoldin target protein actin-related protein 1. Actin-related proteins 2 and 3 have related motifs, but these have altered charge properties. The latter two proteins do not bind prefoldin, although we identify them here as target proteins for the cytosolic chaperonin. Actin fragments containing the two prefoldin interaction regions compete efficiently with actin for prefoldin binding. In addition, they also compete with tubulins, suggesting that these target proteins contact similar prefoldin subunits.

Effect of maturation and aging on beta-adrenergic signal transduction in rat kidney and liver.

The characteristics of the beta-adrenergic signal transduction system were analyzed in kidney and liver membrane preparations from neonatal (2-3 days), mature (2 months), and old (2 years) rats. When comparing kidneys from adult to neonatal rats, we found a higher beta-receptor density and a higher percentage of beta(1)-receptor subtype, lower immunoreactive G(salpha)-protein, a lower ratio between the high and low molecular weight splice variant of G(salpha), lower immunoreactive G(ialpha)-protein, and lower basal adenylate cyclase activity. When comparing livers from adult to neonatal rats, we found lower beta-receptor density and basal adenylate cyclase activity. Very few differences could be detected when comparing mature to old kidneys or livers. Stimulated adenosine 3',5'-cyclic monophosphate (cAMP) synthesis was tissue- and age-dependent. In liver, G-protein- and beta-receptor-stimulated cAMP synthesis mirrored basal adenylate cyclase activity and was highest in liver from neonatal animals. In contrast, cAMP synthesis was significantly more stimulated in kidneys from mature animals than from neonatal and senescent rats. We conclude that: (i) the stoichiometry of the components within the beta-receptor/G-protein/adenylate cyclase complex is not fixed but is both tissue- and age-dependent; (ii) adenylate cyclase enzyme activity is possibly but not necessarily the rate-limiting step in the beta-receptor-mediated synthesis of cAMP; and (iii) there is in vivo evidence for a preferential co-expression of the large splice variant of the G(s)-protein and beta(2)-receptor subtype. It is speculated that this could have important physiological consequences for the development of the kidney.

Activation of phospholipase C by cholecystokinin receptor subtypes with different G-protein-coupling specificities in hormone-secreting pancreatic cell lines.

Phospholipase C (PLC) activity was investigated by stimulation of membrane preparations obtained from insulin (beta-TC3)-, somatostatin (Rin 1027-B2)-, and glucagon (INR1-G9)-producing pancreatic cell lines using the non-hydrolyzable GTP analogue GTPgammaS alone, the C-terminal octapeptide cholecystokinin (CCK-8), or gastrin. All compounds caused a significant 2- to 4.4-fold stimulation of PLC activity in the different cell lines, which was diminished by the non-hydrolyzable GDP analogue GDPbetaS. CCK receptor subtypes were characterized by radioligand binding experiments. High-affinity binding sites for tritiated CCK(A) receptor antagonist L-364,718 (K(d) = 0.24 nM) and tritiated CCK(B) receptor antagonist L-365,260 (K(d) = 0.13 nM) were only present in Rin 1027-B2 cells. High-affinity binding sites for both ligands were not found in beta-TC3 or INR1-G9 cells. Competition binding experiments with non-labeled CCK receptor antagonists CR 1505 (CCK(A) receptor-selective) and CR 2945 (CCK(B) receptor-selective), as well as microphysiometry experiments, resulted in the same receptor distribution. Reverse transcriptase-polymerase chain reaction confirmed the CCK receptor distribution pattern for Rin 1027-B2 cells, but in addition showed the existence of CCK(B) receptors in beta-TC3 cells. Immunoblocking experiments with C-terminal antibodies against different G-protein alpha-subunits demonstrated inhibition of CCK-stimulated PLC activity in beta-TC3 cells by G(q/11)alpha antiserum (70%), in Rin 1027-B2 cells by G(q/11)alpha antiserum (70%) and G(i)-3alpha antiserum (23%), and in INR1-G9 cells by G(q/11)alpha antiserum (60%) and G(o)alpha antiserum (45%). We conclude that CCK receptor subtypes with different G-protein-coupling specificities to PLC are present in the different hormone-secreting cells of the endocrine pancreas.

G-proteins in kidneys of spontaneously hypertensive rats.

G(s alpha)-, total G(i alpha)- and G(q/11alpha)-protein concentrations were investigated by quantitative immunoblotting in membranes of total kidney, renal cortex and medulla as well as in cortical tubules and glomeruli of Spontaneously Hypertensive Rats (SHR) and normotensive Wistar Kyoto rats (WKY), aged 5 weeks, 3 or 8 months. We found that total kidney of 5 week old SHR possess less G(s alpha)-, G(i alpha)- and G(q/11alpha)-proteins than controls. For G(s alpha)-proteins, differences found in total kidney were mirrored both in cortex (tubules and glomeruli) and in medulla. Decreased G(i alpha)-concentrations were accompanied by lower tubular but higher glomerular levels, while medullar levels were also increased. Decreased G(q/11alpha)-concentrations were reflected in decreased glomerular and medullary concentrations. Kidneys of 3 month old SHR and WKY possessed similar concentrations of all G(alpha)-species. In 8 month old SHR similar G(i alpha)-, but decreased G(s alpha)-and G(q/11alpha)-concentrations were observed. The G(s alpha)-decrease was reflected in cortex and medulla, the G(q/11alpha)-decrease in the medulla. We conclude that the main strain-related differences in G(alpha)-concentrations are seen in prehypertensive SHR.

Subcellular localization of neuronal nitric oxide synthase in rat small intestine.

The subcellular localization of neuronal nitric oxide synthase (NOS I, EC 1.14.13.39) was investigated in the longitudinal muscle/myenteric plexus (LM/MP) preparation of rat small intestine. The presence of NOS I, inducible nitric oxide synthase (NOS II), and endothelial nitric oxide synthase (NOS III) was assessed after homogenization and low-speed centrifugation in a postnuclear supernatant by immunological detection after PAGE and Western blotting. Only NOS I was clearly present, whereas NOS II and NOS III were below detection limits. After high-speed centrifugation of the postnuclear supernatant, soluble and particulate fractions were obtained, and the presence of NOS I in these fractions was investigated by measurement of NOS I immunoreactivity and enzyme activity. We found that 90 +/- 1% of NOS I immunoreactivity and 97 +/- 1% of NOS enzyme activity were confined to the soluble fraction of the tissue. Further immunological analysis demonstrated that washing the particulate fraction revealed detectable amounts of NOS I only after concentration of the washing supernatant. Most particulate NOS I remained in the pellet and therefore represents cell organelle-associated enzyme. No NOS I immunoreactivity could be detected as a soluble protein within organelles of the cell. Particulate NOS I could in part be solubilized by Triton X-100 treatment, and the detection of Triton X-100-soluble NOS I was dependent on the antibody used. In conclusion, our results indicate that NOS I in the LM/MP preparation of rat small intestine is mainly soluble and that the particulate NOS I is partly an intrinsic membrane protein and can partly be solubilized by detergent treatment.

Properties of the ventricular adrenergic signal transduction system during ontogeny of spontaneous hypertension in rats.

The purpose of this study was to characterize adrenergic receptors and associated G proteins in ventricles of spontaneously hypertensive rats (SHRs) at different stages of development. The beta- and alpha1-adrenoceptor densities and subtype distribution, and beta-adrenoceptor-G protein coupling were studied by radioligand binding, and levels of G(Salpha), G(ialpha), and G(q/11alpha) protein species were determined by Western blotting in SHRs and Wistar-Kyoto (WKY) control rats aged 3.5 weeks, 3 months, and 8 months. In 3.5-week-old SHRs, both the beta-adrenoceptor density and the percentage of agonist high-affinity binding sites were higher than in age-matched WKY rats. The beta1/beta2-subtype distribution, the alpha1-adrenoceptor density, and the alpha1B/alpha1A-subtype distribution were similar in rats of both strains at all ages. Although essentially no differences in G(salpha) levels between SHRs and WKY rats were detected, higher G(ialpha) and lower Gq/1alpha concentrations were found in 3.5-week-old SHRs. In 3-month-old SHRs, increased levels of Gq/11alpha proteins were observed. In 8-month-old SHRs, none of the parameters was different from those of controls. We conclude that the differences in properties of the adrenergic signal transduction system between SHRs and WKY rats are exclusively observable before and at the onset of the overt hypertension. Moreover, the hypertensive genotype apparently affects G proteins more readily than adrenoceptors.

Changes in the expression of alpha(2)-adrenergic receptor subtypes during maturation of neuronal cells from fetal pig superior cervical ganglia.

The expression of presynaptic alpha(2)-adrenergic receptor (alpha(2)-AR) subtypes was investigated in cultured neurons from fetal pig superior cervical ganglion (SCG). Cells were incubated with chicken antibodies against alpha(2)A-, alpha(2)B- or alpha(2)C-AR subtypes either alone or together with antibodies against dopamine-beta-hydroxylase (DbetaH, a marker for adrenergic neurons) or against choline acetyl transferase (ChAT, a marker for cholinergic neurons). We found immunoreactivity for all three alpha(2)-AR subtypes in SCG-cells when cultured for 8-11 days. The relative expression of the alpha(2)A-subtype was approximately 1/3 of that of alpha(2)B- and alpha(2)C-AR. Co-localisation of all three alpha(2)-AR subtypes was observed in cells expressing DbetaH or ChAT. Increasing the potassium concentration in the culture medium increased the expression of DbetaH and decreased the expression of the alpha(2)A- and alpha(2)C-subtype without altering the expression of the alpha(2)B-subtype. Co-culture of neurons with pig splenocytes enhanced the expression of ChAT and decreased the expression of the alpha(2)B-subtype without altering the expression of alpha(2)A- and alpha(2)C-subtypes. Our results indicate that the three alpha(2)-receptor subtypes are expressed on both noradrenergic and cholinergic nerves. Induction of the noradrenergic phenotype favours the expression of the alpha(2)B-subtype over that of the alpha(2)A- and alpha(2)C-subtype. Conversely, enhancement of the cholinergic phenotype favours the expression of the alpha(2)A- and alpha(2)C-subtypes over that of the alpha(2)B-subtype. Our results suggest that the alpha(2)B-receptor is preferentially associated with noradrenergic nerve endings.

Comparison of the ligand binding and signaling properties of human dopamine D(2) and D(3) receptors in Chinese hamster ovary cells.

Human dopamine D(2) (hD(2)) and D(3) (hD(3)) receptors were expressed at similar, high expression levels in Chinese hamster ovary (CHO) cells, and their coupling to G proteins and further signal transduction pathways were compared. In competition radioligand-binding experiments, guanosine-5'-O-(3-thio)triphosphate (GTPgammaS) treatment of hD(2S)- or hD(3)-CHO cell membranes induced a rightward shift and steeping of the dopamine inhibition curve. This effect was pronounced for hD(2) receptors and small for hD(3) receptors. Activation of G proteins was investigated in [(35)S]GTPgammaS-binding assays. Dopamine stimulated [(35)S]GTPgammaS binding 330 and 70% over basal levels on hD(2)-CHO and hD(3)-CHO cell membranes, respectively. (+)-7-(Dipropylamino)-5, 6,7,8-tetrahydro-2-naphthalenol and PD128907 were partial agonists for both receptors. Haloperidol, risperidone, raclopride, and nemonapride inhibited dopamine-stimulated [(35)S]GTPgammaS binding with potencies comparable to their binding affinities for hD(2) and hD(3) receptors in CHO cell membranes; inverse agonism could not be detected with this assay. Receptor stimulation by dopamine inhibited forskolin-induced cyclic AMP formation in hD(2)-CHO and hD(3)-CHO cells by 70%. Furthermore, the extracellular acidification rate increased when hD(2)-CHO and hD(3)-CHO cells were stimulated by dopamine; this effect was abolished by pertussis toxin pretreatment. In this study, we could demonstrate clear functional effects at different levels of the signaling cascade of hD(2) and hD(3) receptors in CHO cells when expressed at high levels. High-affinity agonist binding to hD(2) and hD(3) receptors was still present, but effects of receptor-G protein uncoupling at hD(3) receptors were small, indicating that hD(3) receptors maintain relatively high-affinity agonist binding in the absence of G proteins.

Distinction between surmountable and insurmountable selective AT1 receptor antagonists by use of CHO-K1 cells expressing human angiotensin II AT1 receptors.

1. CHO-K1 cells that were stably transfected with the gene for the human AT1 receptor (CHO-AT1 cells) were used for pharmacological studies of non-peptide AT1 receptor antagonists. 2. In the presence of 10 mM LiCl, angiotensin II caused a concentration-dependent and long-lasting increase of inositol phosphates accumulation with an EC50 of 3.4 nM. No angiotensin II responses are seen in wild-type CHO-K1 cells. 3. [3H]-Angiotensin II bound to cell surface AT1 receptors (dissociates under mild acidic conditions) and is subject to rapid internalization. 4. Non-peptide selective AT1 antagonists inhibited the angiotensin II (0.1 microM) induced IP accumulation and the binding of [3H]-angiotensin II (1 nM) with the potency order: candesartan > EXP3174 > irbesartan > losartan. Their potencies are lower in the presence of bovine serum albumin. 5. Preincubation with the insurmountable antagonist candesartan decreased the maximal angiotensin II induced inositol phosphate accumulation up to 94% and, concomitantly, decreased the maximal binding capacity of the cell surface receptors. These inhibitory effects were half-maximal for 0.6 nM candesartan and were attenuated by simultaneous preincubation with 1 microM losartan indicating a syntopic action of both antagonists. 6. Losartan caused a parallel rightward shift of the angiotensin II concentration-response curves and did not affect the maximal binding capacity. EXP3174 (the active metabolite of losartan) and irbesartan showed a mixed-type behavior in both functional and binding studies. 7. Reversal of the inhibitory effect was slower for candesartan as compared with EXP3174 and irbesartan and it was almost instantaneous for losartan, suggesting that the insurmountable nature of selective AT1 receptor antagonists in functional studies was related to their long-lasting inhibition.

Human neuropeptide YY1 receptors exert unequal control of the extracellular acidification rate in different cell lines.

The ability of the human neuropeptide YY1 receptor subtype to increase the extracellular acidification rate in different cell lines was investigated by using the Cytosensor Microphysiometer. In CHO-Y1 cells (Chinese Hamster Ovary cells expressing the cloned human neuropeptide YY1 receptor), neuropeptide Y increased the acidification rate by up to 15% of the basal level with a -Log(EC50) of 7.42. As expected for neuropeptide YY1 receptors, this response was potently inhibited by the neuropeptide YY1-selective non-peptide antagonist BIBP3226 ((R)-N2-(diphenylacetyl)-N-[(4-hydroxy-phenyl)methyl]-D-arginine amide). Its enantiomer BIBP3435 ((S)-N2-(diphenylacetyl)-N-[(4-hydroxy-phenyl)methyl]-D-arginin amide) was less potent. The antagonists themselves did not affect the extracellular acidification rate at concentrations up to 10 microM. In SK-N-MC cells (a neuroblastoma cell line of human origin that expresses the neuropeptide YY1 receptor) no change of the acidification rate could be observed in the presence of neuropeptide Y at concentrations up to 1 microM. For control, the neuropeptide YY1 receptors were also investigated by assessing whole cell radioligand binding and, at the functional level, by assessing their ability to decrease the forskolin-induced accumulation of cAMP. The specific (i.e., neuropeptide Y-displaceable) binding of [3H]neuropeptide Y was to a homogeneous class of high-affinity sites in both SK-N-MC and CHO-Y1 cells. The equilibrium dissociation constants for [3H]neuropeptide Y, the total number of binding sites and the kinetic constants for association and for dissociation were similar. Neuropeptide Y produced a dose-dependent inhibition of forskolin-induced cAMP accumulation in SK-N-MC cells (-log(EC50) = 9.40) but it did not affect cAMP accumulation in CHO-Y1 cells. Non-transfected CHO-K1 cells were used as negative control throughout the study. No binding or response could be observed in these cells. Our data suggest that the signalling mechanisms of neuropeptide YY1 receptors are closely related to the cell type in which they are expressed.

Use of elastase as an internal standard in immunoblotting techniques.

A non-radioactive method for relative, semi-quantitative analysis of immunoblots, based on the use of elastase as internal standard and conventional peroxidase staining was devised and applied to the immunoassay of Gs-proteins in crude membrane preparations of rat kidneys. We found that the coefficients of variation of samples, run within the same experiment or run within different experiments, are reduced to half or a quarter of their original value respectively when corrected for elastase as an internal standard, allowing meaningful comparison of these samples.

Effect of age on beta-receptors, Gs alpha- and Gi alpha- proteins in rat heart.

beta-adrenoceptors, Gs alpha- and Gi alpha-proteins were investigated in a crude plasma membrane preparation from ventricles of young (2-4 months) and senescent (22-24 months) Wistar rats. Receptor density, ligand affinity and beta 1/beta 2-receptor ratio were independent of the age of the rats. The percentage of beta-receptors coupled to G-proteins increased with age. An age-related increase in the level of Gs alpha (124%) was paralleled by an increase in the ratio between the high and low molecular weight form of Gs alpha. The level of Gi alpha-protein almost doubled (170%) upon aging. We conclude that the age-related differences are small at the level of the beta-adrenoceptor molecule, but that the increase in Gi alpha-proteins could be responsible for the age-related reduction in myocardial inotropic and chronotropic responses. Moreover, we suggest that the changes in degree of high affinity coupling between beta-receptor and Gs-protein are possibly linked to alterations in the ratio between the Gs-molecular weight subtypes.

Desensitization of alpha 1-, beta- and glucagon receptors in rat hepatocytes: influence of ageing.

The alpha 1-agonist phenylephrine in hepatocytes from mature and senescent rats, and the beta-agonist isoproterenol in hepatocytes from senescent rats, elicited a time-dependent, homologous desensitization of alpha 1- and beta-receptor-mediated glycogenolysis respectively, which was maximal after 20 min of exposure to the agonists. Glucagon triggered desensitization of the glycogenolytic response to glucagon, isoproterenol and phenylephrine, which was maximal after less than 5 min; this was followed by resensitization of the glucagon response only. After 20 min of treatment with phenylephrine or isoproterenol, no change in cellular distribution of alpha 1- or beta-receptors was noticed, using sucrose gradient centrifugation. After a 1-h exposure to both agonists, a shift of both receptors to a light density fraction was found, reflecting receptor internalization. Neither the rate of functional desensitization, nor the degree of receptor internalization was altered upon ageing. We conclude that functional desensitization and internalization of adrenergic receptors in rat hepatocytes are separate events in time and are largely unaffected by the ageing process. Thus, despite the absence of a beta-receptor-mediated glycogenolytic response in hepatocytes from mature rats, isoproterenol triggers the internalization of beta-receptors.

Thermodynamic analysis of isoproterenol binding to beta-adrenoceptors in rat lung membranes.

The thermodynamic properties of the binding of the beta-adrenoceptor agonist isoproterenol and of the antagonist propranolol to beta-adrenoceptors of rat lung were investigated. We found that in our experimental conditions, the high- and low-affinity binding sites for the agonist displayed different properties: the binding to the high-affinity binding site was entropy-driven with a small increase in enthalpy, while agonist binding to the low-affinity binding site was enthalpy-driven. Binding of isoproterenol in the presence of GTP or its non-hydrolyzable analogue GppNHp, and the binding of propranolol were enthalpy-driven with a small increase in entropy.

Influence of aging on the alpha 1-receptor-mediated glycogenolysis in rat hepatocytes.

The involvement of the Ca(2+)-dependent and Ca(2+)-independent, insulin-sensitive pathway in the alpha 1-receptor-control of glycogenolysis was investigated in hepatocytes from young adult, mature adult, and senescent rats. Upon chelation of extracellular Ca2+, phenylephrine caused a similar increase in glucose output that was potently inhibited by insulin, indicating the presence of both pathways in each age group. From the age-related decreasing sensitivity of the Ca(2+)-dependent pathway toward verapamil and nifedipine, and toward insulin, we suggest that the contribution of Ca(2+)-fluxes in eliciting glycogenolysis through the Ca(2+)-dependent pathway decreases upon aging. Both pathways were inhibited by the protein kinase C (PKC) activator, 4 beta-phorbol 12-myristate 13-acetate (PMA); the inhibitory effect was decreased in hepatocytes from mature adult and senescent rats. In conclusion, our results favor the idea that a Ca(2+)-dependent and a Ca(2+)-independent, insulin-sensitive pathway remain involved throughout the life span. We provided the evidence for an impaired regulatory role of protein kinase C and calcium in hepatocytes from the older age groups.

Species and strain-related differences in the expression and functionality of beta-adrenoceptor subtypes in adipose tissue.

The beta-adrenoceptor subtypes which trigger lipolysis in white adipocytes vary markedly between calf and rats, and even between different rat strains. In calf adipocytes, CGP12177, a potent antagonist for beta 1- and beta 2-adrenoceptors (i.e., "classical beta-adrenoceptors") and a partial agonist for atypical beta-adrenoceptors, did not stimulate lipolysis, but inhibited with high affinity (IC50 = 0.66 nM) the lipolytic response to 10 nM isoproterenol. In adipocytes from both Wistar rats and Sprague-Dawley OFA rats, CGP12177 stimulated lipolysis to almost the same extent as isoproterenol. Low concentrations of CGP12177 (3 nM) inhibited part of the lipolytic response to 10 nM isoproterenol in the Sprague-Dawley OFA rat adipocytes, but not in Wistar rats at all ages tested (2-4 weeks, 2-4 months, 24-26 months). Hence, functional beta-adrenoceptors are only classical in calf adipocytes, only atypical in Wistar rat adipocytes and both classical and atypical in Sprague-Dawley OFA rat adipocytes. Binding experiments were performed with 150 pM [125I]CYP. On calf adipocyte membranes, competition binding curves with CGP12177 displayed one high affinity binding site (IC50 = 4.7 nM), whereas the curves for CGP20712 (beta 1-selective antagonist) and ICI118551 (beta 2-selective antagonist) were biphasic. In agreement with the functional data, these results indicate that only beta 1- and beta 2-adrenoceptors are present in calf adipose tissue. For both rat strains, only half of the displaceable [125I]CYP binding sites displayed high affinity for CGP12177 (IC50 = 6.8 to 7.5 nM), and competition binding studies with CGP20712 and ICI118551 indicated that they represent beta 1- and beta 2-adrenoceptors. The remaining [125I]CYP binding sites possessed an about 50 times lower affinity for CGP12177 (IC50 = 260 to 345 nM). They are likely to represent atypical beta-adrenoceptors. It is concluded that the presence and the physiological relevance of beta-adrenoceptor subtypes in adipose tissue may not only be species-related, but also strain-related.

Influence of aging on the beta- and glucagon-receptor-mediated glycogenolysis in rat hepatocytes.

The influence of aging on beta-receptor and glucagon-receptor control of glycogenolysis was investigated in rat hepatocytes. The beta-receptor-induced glucose output was detectable only in senescent rats, was partly dependent on extracellular Ca2+, and was inhibited by 4 beta-phorbol 12-myristate 13-acetate (PMA), insulin, and the Ca(2+)-antagonists, verapamil and nifedipine. Chelation of extracellular Ca2+ potentiated the effect of nifedipine only. In contrast, glucagon-stimulated glycogenolysis, similar in mature and senescent rats, was independent on extracellular Ca2+ and was unaffected by PMA. Verapamil, in senescent rats only, and nifedipine, in mature and senescent rats, inhibited glucagon-stimulated glucose output only in the presence of Ca2+. Insulin inhibited glucagon-induced glucose output, irrespective of the age of the rat and the presence of Ca2+. We conclude that the beta-receptor component in the adrenergic regulation of glycogenolysis in senescent rats consists of a major Ca(2+)-independent and a minor Ca(2+)-dependent part, displaying different sensitivity towards protein kinase C (PKC), Ca(2+)-antagonists, and insulin. Aging does not change the capacity of glucagon to induce a full glycogenolytic response in the absence of extracellular Ca2+; Ca(2+)-influx, however, seems to be involved when extracellular Ca2+ is present, and this sensitivity is increased on aging.