Quantitative analysis of muscle breakdown during starvation in the marine flatfish Pleuronectes platessa

Summary The present study describes the effects of starvation for a duration of four months on the ultrastructure of skeletal muscles from the marine flatfish (Pleuronectes platessa L.). Starvation is associated with a decrease in resting metabolic rate from 20.1±2.2 to 11.6±1.5mg-O₂/kg/h (P<0.05) and muscle wasting. Median fibre size fell from 700 m² to 500 m² in intermediate (fast oxidative) and from 1,800 m² to 600 m² in starved, white (fast-glycolytic) muscle fibres. In contrast, median fibre size in red (slow oxidative) muscle remained within the range 300–400 m². The fraction of red fibre volume occupied by myofibrils (58.6%) and mitochondria (24.5%) did not change significantly with starvation. There was, however, a decrease in stored lipid (10.7% to 3.2%) and an alteration in the structure of the cristae in mitochondria from red muscle.Atrophy of white muscle fibres is associated with a decrease in both the diameter and fractional volume occupied by myofibrils (85.7% to 61.9% P < 0.01). In a high proportion of white fibres peripheral degeneration of Z-discs is evident causing an unravelling of the thin filament lattice. It is suggested that this allows a partial decrease in myofibril diameter and hence the maintenance of contractile function in muscle from starved fish. In severely degenerating white fibres, disorganised thick and thin filaments and numerous multimembrane lysosome-like vesicles are observed.Starvation results in an increase in the average content of mitochondria in white fibres from 2.2 to 6.7% (P<0.01). In fed plaice mitochondria constitute less than 1% of the volume of the white fibre in 43.5% of the fibres. The proportion of white fibres containing more than 6% mitochondria increases from 6.5% to 58% with starvation.     

The fine structure of red and white myotomal muscle fibres of the coalfish (Gadus virens)

Summary The fine structure of the red and white myotomal muscles of a marine teleost, the coalfish Gadus virens, has been examined and ultrastructural measurements and analyses carried out. The sarcomere lengths of the red and white fibres were found to be 1.60 minimum, 1.82 maximum and 1.70 minimum, 1.85 maximum, respectively. No significant difference was found between the red and white fibres in their percentage of sarcoplasmic reticulum and T system. Both were found to have regularly occurring triads at the Z disk level, to have distinctive M lines and to be multiply innervated. Ultrastructurally the two fibres can be distinguished by the thicker Z line and more abundant mitochondria of the red fibre, and by the ribbon-shaped peripheral myofibrils of the white fibres. The structure of the fibres in these two types of muscle is discussed in relation to their possible role in swimming.This work was supported by a research grant from the National Environmental Research Council.     

Structural and histochemical characterization of the cotyledon storage organelles of jojoba (Simmondsia chinensis)

Summary Jojoba cotyledon cells are isodiametric and 44 m in diameter. They contain two prominent storage organelles—wax bodies and protein bodies. Isolated and intact wax bodies react positively with several lipid histochemical stains; they are spherical and 1.5 m in diameter. Protein bodies are irregular in shape, but usually appear spherical. They range in diameter from 7–15 m, are unit membrane bound, have an irregular grainy matrix and do not contain a globoid inclusion. Protein bodies stain positively for both protein and lipid. The possible composition and role of this protein/lipid body complex is discussed.     

„Rote“ Muskelfasern

Zusammenfassung Die Fasern des roten und langsamen M. soleus von Ratte, Kaninchen und Katze und des roten, jedoch schnellen, M. vocalis des Kaninchens wurden licht- und elektronenmikroskopisch untersucht und mit den verschiedenen Fasertypen aus dem M. tibialis anterior der Ratte und dem M. gastrocnemius des Kaninchens und der Katze verglichen. M. soleus und M. vocalis (einschließlich M. thyreoarytenoideus) enthalten nur einen mitochondrienreichen Fasertyp. Im schnellen M. vocalis ist der Z-Streifen schmal (50–60 nm), das sarcoplasmatische Reticulum ist gut entwickelt. Die Anordnung von Reticulum und Mitochondrion ist ähnlich wie in Herzmuskelzellen. Wie auch in anderen langsamen Muskeln verschiedener Tiere ist im M. soleus der Z-Streifen breit (100–120 nm), Triaden und Reticulum sind selten, und die Filamente bilden unregelmäßige Areale anstelle von Fibrillen. Hierin gleichen die Fasern des M. soleus den (mitochondrienreichen) C-Fasern eines entsprechenden gemischten Muskels; dagegen zeigen die Zwischentyp-(B-)Fasern schmale Z-Linien (50–70 nm), isodiametrische Fibrillen und mehr Triaden als die C-Fasern. Entgegen der bisherigen Vermutung, die auf der histochemischen Zuordnung der SoleusFasern zum Typ B und der Vocalis-Fasern zum Typ C beruht, ist daher anzunehmen, daß die langsamen motorischen Einheiten eines gemischten Muskels aus C- und nicht aus B-Fasern bestehen.In einigen Muskeln sind die Sarcomere der C-Fasern länger als die der B-(und A-) Fasern. Im M. tibialis anterior der Ratte verschwindet der Unterschied von 8,5% bei 2,6 m Sarcomerlänge bei der Dehnung auf 2,8 m mittlere Sarcomerlänge; vermutlich weil die Ruhedehnungskurve zunehmend steiler wird. Die isometrische Extraspannung im Tetanus ist bei 120% der Ruhelänge, d.h. bei 2,7 m Sarcomerlänge. am größten. Daher muß bei 2,6 m mittlerer Sarcomerlänge die Kraft der C-Fasern die der B-Fasern übertreffen.Rote Muskeln sind besser vaskularisiert als weiße Muskeln. Für die Mm. soleus und gastrocnemius der Katze verhalten sich die Kapillardichten (Kapillaren/mm² Muskelfaserquerschnitt) wie 2,7:1. Dieser Wert entspricht dem Verhältnis zwischen den Größen für die Durchblutung (ml/min × 100 g) in Ruhe und bei maximaler Gefäßerweiterung.

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Redmuscle fibres

Summary Muscle fibres of the red and slow contracting soleus of rat, rabbit and cat and of the red however fast contracting thyreoarytenoid of rabbit are compared with different fibre types in the anterior tibial muscle of rat and in the gastrocnemius of rabbit and cat. With respect to fibre types soleus and thyreoarytenoid (including m. vocalis) are homogeneous and both being rich in mitochondria. The fast thyreoarytenoid shows a narrow Z-line (50–60 nm) and a well developed sarcoplasmic reticulum. The pattern of reticulum and mitochondria resembles more that of heart muscle cells than of skeletal muscle fibres. Like many slow contracting muscles of different animals the soleus fibres display a wide Z-line (100–120 nm), few triads, little reticulum and irregularly shaped areas of myofilaments instead of fibrils. In that soleus fibres equal fibres of type C (rich in mitochondria) in a corresponding heterogeneous muscle, whereas intermediate (type B) fibres reveal narrow Z-lines (50–70 nm), isodiametrically shaped myofibrils and more triads than C-fibres. Therefore it is far more likely that the slow motor units of a mixed muscle consist of C-fibres than of B-fibres. This is at variance with the histochemical designation of soleus fibres as type B and thyreoarytenoid fibres as type C.In some muscles in C-fibres the sarcomeres are longer than in B-(and A-)fibres. In the anterior tibial muscle of rat this difference is 8.5% at a mean sarcomere length of 2.6 m, and disappears at a mean length of 2.8 m, probably due to the steeper slope of the length tension diagram at rest. Since the isometric extratension in a tetanus is highest at 120% resting length (corresponding to about 2.7 m sarcomere length), the force of C-fibres exceeds that of B-fibres at 2.6 m but not at 2.8 m sarcomere length.Red and white muscle differ with respect to vascularisation. The relation between the densities of capillaries in soleus and gastrocnemius of cat is 2.7:1 and equals the relation between the blood flows through these muscles during rest and maximum vasodilatation.

Herrn Prof. Dr. Bargmann nachträglich zum 65. Geburtstag.     

Fine structure and metabolic adaptation of red and white muscles in tuna

Synopsis An electron microscopic study of the red and white muscle fibres in the trunk musculature of the Kawakawa tuna (Euthynnus affinis) was carried out with a view to correlating their structure with metabolic adaptation. The red fibres which are considerably smaller in diameter (34.58 m ± 6.16 S.D.) are characterized by their high content of myoglobin, mitochondria, lipid droplets and glycogen granules. The white fibres which are relatively larger in diameter (66.03 m ± 11.59 S.D.) are characterized by their lack of myoglobin, low mitochondria) density, high content of glycogen granules and the conspicuous absence of lipid droplets. The characteristics in fine structure of the two fibre types are discussed in the light of their metabolic adaptation, the red fibres as being adapted for long term cruising movement utilizing lipid as the main source of energy and the white fibres for short bursts of activity metabolizing glycogen as the chief fuel.The tuna, with the acquisition of the counter-current heat exchange system which provides for the retention of the heat generated from high substrate oxidation in the red muscle and an efficient respiratory system, it is postulated, is well adapted for high speed sustained swimming.     

Muscle fibre types and metabolism in post-larval and adult stages of notothenioid fish

Summary A histochemical study was carried out on muscle fibre types in the myotomes of post-larval and adult stages of seven species of notothenioid fish. There was little interspecific variation in the distribution of muscle fibre types in post-larvae. Slow fibres (diameter range 15–60 m) which stained darkly for succinic dehydrogenase activity (SDHase) formed a superficial layer 1–2 fibres thick around the entire lateral surface of the trunk. In all species a narrow band of very small diameter fibres (diameter range 5–62 m), with only weak staining activity, occurred between the skin and slow fibre layer. These have the characteristics of tonic fibres found in other teleosts. The remainder of the myotome was composed of fast muscle fibres (diameter range 9–75 m), which stain weakly for SDHase, -glycerophosphate dehydrogenase, glycogen and lipid. Slow muscle fibres were only a minor component of the trunk muscles of adult stages of the pelagic species Champsocephalus gunnari and Pseudochaenichthys georgianus, consistent with a reliance on pectoral fin swimming during sustained activity. Of the other species examined only Psilodraco breviceps and Notothenia gibberifrons had more than a few percent of slow muscle in the trunk (20%–30% in posterior myotomes), suggesting a greater involvement of sub-carangiform swimming at cruising speeds. The ultrastructure of slow fibres from the pectoral fin adductor and myotomal muscles of a haemoglobinless (P. georgianus) and red-blooded species (P. breviceps), both active swimmers, were compared. Fibres contained loosely packed, and regularly shaped myofibrils numerous mitochondria, glycogen granules and occasional lipid droplets. Mitochondria occupied >50% of fibre volume in the haemoglobinless species P. georgianus, each myofibril was surrounded by one or more mitochondria with densely packed cristae. No significant differences, however, were found in mean diameter between fibres from red-blooded and haemoglobinless species. The activities of key enzymes of energy metabolism were determined in the slow (pectoral) and fast (myotomal) muscles of N. gibberifrons. In contrast to other demersal Antarctic fish examined, much higher glycolytic activities were found in fast muscle fibres, probably reflecting greater endurance during burst swimming.     

The histochemical demonstration of myoglobin and succinic dehydrogenase activity in the tibialis anterior muscle of the rabbit

Summary The staining reactions for myoglobin and succinic dehydrogenase activity in the tibialis anterior of the rabbit demonstrate four types of muscle fibre. These may be distinguished by their intensity of staining for myoglobin and the distribution of the mitochondria shown by the dehydrogenase reaction.The large fibres (70–80 m diameter) which contain many mitochondria evenly scattered throughout the fibre contain much myoglobin. Smaller fibres (45–60 m diameter) which show an identical staining reaction for the dehydrogenase reaction contain less myoglobin. This suggests that myoglobin may be present to aid the diffusion of oxygen into muscle fibres.     

Architecture of the media of the arterial vessels in the dog brain: A scanning electron-microscopic study

Summary The architecture of the media of arterial vessels in dog brain was investigated using scanning electron microscopy. The arrangement and shape of the circularly-oriented smooth muscle cells varied with vessel diameter: The arteries (>100 m in diameter) had 4–10 layers of spindle-shaped smooth muscle cells; the muscular arterioles (30–100 m), 2–3 layers of spindle-shaped smooth muscle cells; the terminal arterioles (10–30 m), a compact layer of spindle-shaped smooth muscle cells with more dominant nodular or rod-like processes and thin lateral processes; and the precapillary arterioles (5–15 m), a less compact layer of branched smooth muscle cells.Longitudinally-oriented muscles were observed in the medio-adventitial border. The distribution and arrangement of these muscles varied with vessel size: in the large arteries (> 300 m in diameter), at the branching sites only; in the small arteries (100–300 m), at both the branching and non-branching sites; in the muscular arterioles, at both the branching and non-branching sites in a reticular arrangement with some muscle cells having an asteroid appearance; in the terminal aterioles, only asteroid-like muscle cells were found at the branching and non-branching sites.     

Three-dimensional arrangement of sarcoplasmic reticulum in crayfish as seen by high voltage electron microscope

Summary Sections several m thick of single fibres from the crayfish muscle were examined by means of a high voltage electron microscope to find out the geometry of the sarcoplasmic reticulum (SR). A sufficient contrast of the SR was achieved by lead acetate histochemical method for calcium. Longitudinally oriented files of SR vesicles at the level of A bands and interruptions in otherwise continuous SR net are the most conspicuous features.     

Metabolic variability within individual fibres of the cat tibialis posterior and diaphragm muscles

Summary Variance in succinate dehydrogenase activity along the transverse and longitudinal axes of fibres from the cat tibialis posterior and diaphragm muscles was determined in order to estimate the three-dimensional distribution of mitochondria within single fibres. The variance (coefficient of variation) in succinate dehydrogenase activity along the transverse fibre axis was greatest in type IIB fibres from both muscles. Intracellular compartmentalization (i.e. subsarcolemmal vs central core differences in succinate dehydrogenase activity) was observed only in type II fibres from the tibialis posterior; the succinate dehydrogenase activity of the subsarcolemmal region was significantly greater than that of the central core. The extent of succinate dehydrogenase variance along the longitudinal fibre axis was dependent on the total length of the fibre segment analyzed, the muscle, and fibre type. The coefficient variation for short fibre segments, i.e. 40 m, was significantly lower than that for much longer fibre segments (840 m). Significant differences in the coefficient variation for 840 m fibre segments were observed between the diaphragm and tibialis posterior muscles. The longitudinal variance in succinate dehydrogenase activity was higher in diaphragm muscle fibres. The succinate dehydrogenase variance along the longitudinal axis of type II fibres was significantly greater than that of the type I fibre population. These results indicate that mitochondria are heterogeneously distributed within muscle fibres. Possible functional implications of such intrafibre metabolic variance are discussed.     

A morphometric analysis of regional differences in myotomal muscle ultrastructure in the juvenile eel (Anguilla anguilla L.)

Summary Subpopulations of fast and slow fibres within the trunk musculature of elvers were examined using morphometric analysis of electron micrographs. Fibre regions were characterised by their histochemical staining characteristics, and individual fibres located using a coordinate mapping system utilising morphological features as reference points. Percentages of fibre volume occupied by mitochondria, myofibrils, sarcoplasmic reticulum (S.R.), and T-system were determined in each of the fibre groups, along a transect from the skin to the vertebral column (fibres 1–14, respectively).The fine structure of slow (red) fibres (1–2 fibres deep) is relatively homogeneous throughout its range, giving mean values for mitochondria, 21.4%; myofibrils, 61.0%; S.R., 2.10%; T-system, 0.31%. The fibres are relatively small (204 m²) and the mitochondrial cristae poorly developed.In contrast, there is a marked heterogeneity in the ultrastructure of fast (white) fibres, dependent on both position and size. The moderately small (333 m²) superficial fast fibres (3–4 fibres deep) have a significantly higher mitochondrial content (7.6%) than the larger deep fibres (1.2%) (6–12 fibres deep, 775 m²). The mean fractional volumes occupied by myofibrils, S.R., and T-system in the deep fibres are: 80.4%, 5.95%, and 0.38%, respectively. Fibres < 100 m² constitute up to 5% of the fast muscle and have a significantly higher mitochondrial volume (4.3%), more glycogen granules, and a slightly lower volume of S.R. (5.57%) than larger fibres.It is suggested that metabolic subpopulations of fast fibres correspond to different stages of fibre growth. The relatively poorly developed S.R. of eel fast muscle is thought to be correlated with the low frequency, high amplitude nature of the propagated waveform found in anguilliform locomotion.     

Karyometry of nuclei in liver cells

Summary In untreated adult male albino rats nuclear volume and the percentage of binucleate cells were determined in the first layer of hepatocytes adjacent to hepatic venous branches of varying diameters (<40 m, 40 m–80 m, 80 m–120m, 120 m–160 m, >160 m), and in the third and fourth layer of hepatocytes in the remainder of the perivenous parenchyma. In the first layer of hepatocytes adjacent to the vascular structures means of nuclear volume are significantly lower and percentage of binucleate cells significantly higher than in the cells of the remainder of the perivenous parenchyma. Within each area measured distribution curves of nuclear volume classes were homogeneous but showed heterogeneity in comparison with each other. The morphometric data presented in this study strongly support the opinion of the heterogeneity of liver cells in the perivenous zone, as previously postulated on the basis of histochemical investigations.Supported by the Deutsche Forschungsgemeinschaft (Hi 318/2-1)     

Immunocytochemical study of the hypothalamo-neurohypophysial system

Summary An antiserum cross-reactive against ovine neurophysins-I-II and -III has been used in conjunction with the immunoperoxidase histochemical procedure to localize the cells of the sheep paraventricular (PVN) and supraoptic nuclei (SON). In order to describe the topographical distribution of the SON and PVN a study was made on the serial sections cut (a) transversely from rostral to caudal positions and (b) sagittally from lateral to medial positions of the hypothalamus.The cells of the SON, when examined in the transverse aspect, extended approximately 1900 caudally and when examined in the sagittal plane were contained within a lateral-medial distance of 4830 . In each case the SON cells lay adjacent to the optic chiasm.As sections were cut transversely, the cells of the PVN first appeared in a rostral position defined as 0 and close to the ventral lining of the third ventricle. This general ventral and ventro-lateral distribution of cells maintained up to a caudal distance of approximately 840 . From positions 1260–2310 there was a dramatic dorsal shift of the PVN cells which by this time had also extended laterally. The total rostral-caudal distance occupied by the PVN cells was 3150 . That the lateral-medial distance occupied by the PVN was small (1050 ) was determined on examining the magnocellular nuclei in sagittal section.This work was financed by a grant awarded by the Medical Research Council of New Zealand     

The spermatozoon of Oikopleura dioica Fol (Larvacea,Tunicata)

Summary The spermatozoon of Oikopleura dioica is about 30 m long, with a spherical head, about 1 m wide, a 3 m long and 1 m wide midpiece, and a 25 m long tail with a tapered end piece. The head contains a nucleus with the chromatin volume limited to about 0.1 m³. A small acrosome is found in an anterior inpocketing, and a flagellar basal body in a posterior inpocketing of the nucleus. The midpiece contains a single mitochondrion with the flagellar axoneme embedded in a groove along its medial surface. The flagellar axoneme has the typical 9 + 2 substructure, and the basal body the typical 9+0 substructure. A second centriole and special anchoring fibres are absent.     

Myelogenesis and estimation of the number of axons in the anterior commissure of the chick (Gallus gallus)

Summary By use of light- and electron microscopy the anterior commissure of the chick was studied at different times during development. Between the 19th day of incubation and the 35th day after hatching the cross-sectional area of the anterior commissure, as determined from mid-sagittal sections, undergoes a 6-fold increase in size. Thereafter the area remains fairly constant. The total number of fibres in the anterior commissure was estimated to be 89000. The full complement of fibres is already present by the 19th day of incubation. Myelogenesis occurs mainly between the 19th day of incubation and the 35th day after hatching, concomitant with the increase in cross-sectional area. From the 35th day after hatching, myelinated fibres comprise approximately 40% of the total number of fibres. The median diameter of unmyelinated fibres is about 0.35–0.40 m. The median diameters of myelinated axons and fibres are 0.8–1.0 m and 1.1–1.3 m, respectively.     

Cholinergic nerves in the human liver

Summary The cholinergic innervation of the human liver was studied. Slices (150–200m thick) of human liver and of the greater hepatic blood vessels (hepatic artery and vein, portal vein) were incubated in a solution of 6-hydroxydopamine (6-HDA) in order to obtain a selective degeneration of adrenergic nerves. Controls were prepared from samples incubated with buffer alone. The slices were cut on a cryostat into 15–20m thick sections and processed for the histochemical detection of cholinesterases.Cholinergic nerve fibres innervate the extra hepatic and the intrahepatic branches of the hepatic artery, the portal vein as well as the hepatic vein. Fewer cholinergic fibres innervate the hepatocytes and the hepatic sinusoids. The 6-HDA treatment does not seem to alter the pattern of the cholinergic innervation of the liver. The findings indicate the presence of a cholinergic parasympathetic innervation in the human liver.     

Clonal propagation of Callistemon by tissue culture techniques

Callus development in Callistemon viminalis was readily achieved when axillary buds derived from nodal tissue were placed in a medium containing macro- and micro-nutrients, sucrose (0.06 M), inositol (300 M), nicotinic acid (20 M), pyridoxine hydrochloride (3 M), thiamine hydrochloride (2 M), riboflavin (10 M), cytokinins (5 M) and auxins (0.1 M). The presence of benzylaminopurine (5 M) and p-chlorophenoxyacetic acid (0.1 M) promoted the most vigorous callus development and sprout formation. Rooting of nodal material was rare but occurred readily following the transference of sprouts developed on callus to a basal medium containing sucrose and salts. Root initiation was stimulated, however, by the presence of auxins. Chlorophenoxyacetic acid while stimulating root initiation repressed root growth. Indole butyric acid stimulated both root initiation and shoot growth at concentrations of 0.005 to 0.1 M. The treatment of choice for rooting and shoot growth was the addition of indole butyric acid at a concentration of 0.01 M.     

Leptomeric fibrils and T-tubule desmosomes in the Z-band region of the mouse heart papillary muscle

Summary The papillary muscle of the heart of adult white mice is investigated. Intrafibrillarly located leptomeric fibrils, frequently encountered in the Z-band region of the myofibrils. The leptomeric fibrils are always running in a transverse direction and often in close proximity to the transverse tubules (which are also located at this level). There seems to be a close connection between the dense striae of the leptofibrils and the Z-bands of ordinary myofibrils. The leptomeric fibrils are spindle-shaped and have a length varying between 0.6 and 1.2 m. The banding periodicity of the fibrils is approximately 0.16 m.Occasionally desmosomes are observed in the T-tubule system.     

Further histochemical properties of rabbit skeletal muscle fibres

Summary The histochemical activities of succinic dehydrogenase (SDH), creatine kinase (CK), sarcoplasmic reticular ATPase (SR-ATPase) and myosin ATPase were studied in serial sections of rabbit adductor muscle. Three fibre types were distinguished depending upon the distribution of the enzyme activities. The type II white fibres posessing minimal SDH showed high myosin ATPase, SR-ATPase and ATPase dependent CK activities. Red oxidative fibres showing high SDH fell into two distinct groups: One category had mainly a peripheral localization of SDH and showed an enzymatic profile identical to that of type II white fibres. The second category of red fibres displayed both a homogeneous distribution of small diformazan granules throughout the fibre as well as a sub-sarcolemmal collection when tested for SDH activity but possessed very low amounts of reaction product of the various enzymes of the energetic metabolism studied. Since it is well established that the myosin ATPase of a fibre correlates with its contraction time, the present histochemical investigation provides further support for this concept by demonstrating the presence of high SR-ATPase and ATPase dependent CK activities in all white and red fibres rich in myosin ATPase.     

Ultrastructure of arterioles in the cat brain

Summary A total of 110 arterioles were examined in the brains of cats; different sites were studied including the cortex, putamen, pons and crus cerebri. No internal elastic laminae were seen in the subendothelial space, although occasional fragments of elastic material were present in the larger arterioles. The media was composed of one, two or three layers of smooth muscle cells which interlocked in such a way that the vessel wall thickness was constant. Numerous tight junctions were seen between adjacent smooth muscle cells and between the endothelium and smooth muscle cells. Apart from the usual cell organelles, the smooth muscle cells of arterioles had numerous dense patches on the cell surface. The structure of the adventitia varied according to the diameter of the vessel and the site in the brain; it contained adventitial cells, bundles of collagen fibres and nerve fibres. Innervation of arterioles was more constant in the brain stem than in the cortex. Metarterioles had less specialised, atypical smooth muscle cells, a discontinuous media and numerous, extensive myoendothelial tight junctions; they were not innervated by nerve fibres. The diameter of metarterioles was less than 10 m whereas that of arterioles was 10–45 m. The possible functional aspects of arteriolar innervation are discussed.