No abstract

alpha(2)-Adrenergic receptors play an essential role in regulating neurotransmitter release from sympathetic nerves and from adrenergic neurons in the CNS. However, the role of each of the three highly homologous alpha(2)-adrenergic receptor subtypes (alpha(2A), alpha(2B), alpha(2C)) in this process has not been determined unequivocally. To address this question, the regulation of norepinephrine and dopamine release was studied in mice carrying deletions in the genes encoding the three alpha(2)-adrenergic receptor subtypes. Autoradiography and radioligand binding studies showed that alpha(2)-receptor density in alpha(2A)-deficient brains was decreased to 9 +/- 1% of the respective wild-type value, whereas alpha(2)-receptor levels were reduced to 83 +/- 4% in alpha(2C)-deficient mice. These results indicate that approximately 90% of mouse brain alpha(2)-receptors belong to the alpha(2A) subtype and 10% are alpha(2C)-receptors. In isolated brain cortex slices from wild-type mice a non-subtype-selective alpha(2)-receptor agonist inhibited release of [(3)H]norepinephrine by maximally 96%. Similarly, release of [(3)H]dopamine from isolated basal ganglion slices was inhibited by 76% by an alpha(2)-receptor agonist. In alpha(2A)-receptor-deficient mice, the inhibitory effect of the alpha(2)-receptor agonist on norepinephrine and dopamine release was significantly reduced but not abolished. Only in tissues from mice lacking both alpha(2A)- and alpha(2C)-receptors was no alpha(2)-receptor agonist effect on transmitter release observed. The time course of onset of presynaptic inhibition of norepinephrine release was much faster for the alpha(2A)-receptor than for the alpha(2C)-subtype. After prolonged stimulation with norepinephrine, presynaptic alpha(2C)-adrenergic receptors were desensitized. From these data we suggest that two functionally distinct alpha(2)-adrenergic receptor subtypes, alpha(2A) and alpha(2C), operate as presynaptic inhibitory receptors regulating neurotransmitter release in the mouse CNS.

Together these data imply that nmDC phenotypical differ from omDC which might result in diverse functional properties and might be of relevance for selecting routes for immunotherapy of atopic diseases. Moreover these data provide a basis for further studies investigating immunological mechanisms underlying mucosal immunotherapy.

Although G protein-coupled receptor-mediated signaling is one of the best studied biological events, little is known about the kinetics of these processes in intact cells. Experiments with neurons from ␣ 2A -adrenergic receptor knockout mice suggested that the ␣ 2A -receptor subtype inhibits neurotransmitter release with higher speed and at higher action potential frequencies than the ␣ 2C -adrenergic receptor. Here we investigated whether these functional differences between presynaptic ␣ 2 -adrenergic receptor subtypes are the result of distinct signal transduction kinetics of these two receptors and their coupling to G proteins. ␣ 2A -and ␣ 2C -receptors were stably expressed in HEK293 cells at moderate (ϳ2 pmol/mg) or high (17-24 pmol/mg) levels. Activation of G protein-activated inwardly rectifying K ؉ (GIRK) channels was similar in extent and kinetics for ␣ 2A -and ␣ 2C -receptors at both expression levels. However, the two receptors differed significantly in their deactivation kinetics after removal of the agonist norepinephrine. ␣ 2C -Receptor-activated GIRK currents returned much more slowly to base line than did ␣ 2A -stimulated currents. This observation correlated with a higher affinity of norepinephrine at the murine ␣ 2C -than at the ␣ 2A -receptor subtype and may explain why ␣ 2C -adrenergic receptors are especially suited to control sympathetic neurotransmission at low action potential frequencies in contrast to the ␣ 2A -receptor subtype. G protein-coupled receptors (GPCRs)1 transfer a large diversity of extracellular signals into the cell interior, including light, neurotransmitters, and hormones. Although GPCRs represent some of the best studied signaling molecules, relatively little information exists about the kinetics of signal transduction by these receptors (except for rhodopsin) in intact cells. However, more detailed knowledge about the kinetic properties of GPCR signal transduction would be of particular interest to determine the physiological significance of closely related receptor subtypes, which can be activated by the same endogenous agonist but differ in their biological function.Functional data on ␣ 2 -adrenergic receptor subtypes suggest that they differ in their signaling kinetics. Interestingly, several physiological differences were identified between presynaptic ␣ 2A -and ␣ 2C -receptor subtypes (1). In mouse atria, the ␣ 2A -subtype inhibited norepinephrine release at high stimulation frequencies whereas the ␣ 2C -receptor operated at lower levels of sympathetic nerve activity (1). Moreover, inhibition of norepinephrine release mediated by the ␣ 2A -subtype occurred much faster than inhibition by the ␣ 2C -receptor. These findings indicate that two presynaptic receptors in the inhibitory feedback loop of transmitter release may differentially regulate synaptic transmission. Several explanations may account for these functional differences. ␣ 2 -Adrenergic receptor subtypes have been shown to differ in their signal transduction, agonistdependent internalization and receptor...

The loss of cartilage and bone because of congential defects, trauma and after tumor resection is a major clinical problem in head and neck surgery. The most prevalent methods of tissue repair are through autologous grafting or using implants. Tissue engineering applies the principles of engineering and life sciences in order to create bioartificial cartilage and bone. Most strategies for cartilage tissue engineering are based on resorbable biomaterials as temporary scaffolds for chondrocytes or precursor cells. Clinical application of tissue-engineered cartilage for reconstructive head and neck surgery as opposed to orthopedic applications has not been well established. While in orthopedic and trauma surgery engineered constructs or autologous chondrocytes are placed in the immunoprivileged region of joints, the subcutaneous transplant site in the head and neck can lead to strong inflammatory reactions and resorption of the bioartificial cartilage. Encapsulation of the engineered cartilage and modulation of the local immune response are potential strategies to overcome these limitations. In bone tissue engineering the combination of osteoconductive matrices, osteoinductive proteins such as bone morphogenetic proteins and osteogenic progenitor cells from the bone marrow or osteoblasts from bone biopsies offer a variety of tools for bone reconstruction in the craniofacial area. The utility of each technique is site dependent. Osteoconductive approaches are limited in that they merely create a favorable environment for bone formation, but do not play an active role in the recruitment of cells to the defect. Delivery of inductive signals from a scaffold can incite cells to migrate into a defect and control the progression of bone formation. Rapid osteoid matrix production in the defect site is best accomplished by using osteoblasts or progenitor cells.

Cartilage tissue engineering holds considerable promise for orthopaedic and reconstructive head and neck surgery. With an increasingly ageing population, the number of patients affected by arthritis and recurrent joint pain is constantly growing, along with the associated socio-economic costs. In head and neck surgery reconstructive procedures gain increasing importance in multimodal tumour therapies. These procedures require the harvesting of large amounts of donor tissue, which causes significant donor site morbidity. Therefore, in vitro-engineered cartilage may provide for a cost-effective and clinically valuable medical need. This article presents an overview of the clinical background as well as considerations for engineered cartilage in the head and neck, and provides examples of cartilage tissue engineering based on various scaffolds.

Small-vessel disease or cerebral microangiopathy (CMA) is a common finding in elderly people. It is related to a variety of vascular risk factors and may finally lead to subcortical ischemic vascular dementia. Because vessel stiffness is increased, we hypothesized that slow spontaneous oscillations are reduced in cerebral hemodynamics. Accordingly, we examined spontaneous oscillations in the visual cortex of 13 patients suffering from CMA, and compared them with 14 agematched controls. As an imaging method we applied functional near-infrared spectroscopy, because it is particularly sensitive to the microvasculature. Spontaneous low-frequency oscillations (LFOs) (0.07 to 0.12 Hz) were specifically impaired in CMA in contrast to spontaneous very-lowfrequency oscillations (0.01 to 0.05 Hz), which remained unaltered. Vascular reagibility was reduced during visual stimulation. Interestingly, changes were tightly related to neuropsychological deficits, namely executive dysfunction. Vascular alterations had to be attributed mainly to the vascular risk factor arterial hypertension. Further, results suggest that the impairments might be, at least partly, reversed by medical treatment such as angiotensin-converting enzyme inhibitors/angiotensin II receptor blockers. Results indicate that functional near-infrared spectroscopy may detect changes in the microvasculature due to CMA, namely an impairment of spontaneous LFOs, and of vascular reagibility. Hence, CMA accelerates microvascular changes due to aging, leading to impairments of autoregulation.

Tissue engineering is a field of research with interdisciplinary cooperation between clinicians, cell biologists, and materials research scientists. Many medical specialties apply tissue engineering techniques for the development of artificial replacement tissue. Stages of development extend from basic research and preclinical studies to clinical application. Despite numerous established tissue replacement methods in otorhinolaryngology, head and neck surgery, tissue engineering techniques opens up new ways for cell and tissue repair in this medical field. Autologous cartilage still remains the gold standard in plastic reconstructive surgery of the nose and external ear. The limited amount of patient cartilage obtainable for reconstructive head and neck surgery have rendered cartilage one of the most important targets for tissue engineering in head and neck surgery. Although successful in vitro generation of bioartificial cartilage is possible today, these transplants are affected by resorption after implantation into the patient. Replacement of bone in the facial or cranial region may be necessary after tumor resections, traumas, inflammations or in cases of malformations. Tissue engineering of bone could combine the advantages of autologous bone grafts with a minimal requirement for second interventions. Three different approaches are currently available for treating bone defects with the aid of tissue engineering: (1) matrix-based therapy, (2) factor-based therapy, and (3) cell-based therapy. All three treatment strategies can be used either alone or in combination for reconstruction or regeneration of bone. The use of respiratory epithelium generated in vitro is mainly indicated in reconstructive surgery of the trachea and larynx. Bioartificial respiratory epithelium could be used for functionalizing tracheal prostheses as well as direct epithelial coverage for scar prophylaxis after laser surgery of shorter stenoses. Before clinical application animal experiments have to prove feasability and safety of the different experimental protocols. All diseases accompanied by permanently reduced salivation are possible treatment targets for tissue engineering. Radiogenic xerostomia after radiotherapy of malignant head and neck tumors is of particular importance here due to the high number of affected patients. The number of new diseases is estimated to be over 500,000 cases worldwide. Causal treatment options for radiation-induced salivary gland damage are not yet available; thus, various study groups are currently investigating whether cell therapy concepts can be developed with tissue engineering methods. Tissue engineering opens up new ways to generate vital and functional transplants. Various basic problems have still to be solved before clinically applying in vitro fabricated tissue. Only a fraction of all somatic organ-specific cell types can be grown in sufficient amounts in vitro. The inadequate in vitro oxygen and nutrition supply is another limiting factor for the fabrication of complex tissues or organ sys...