Occurrence of FMRF amide-like immunoreactive nerve fibers in guinea pig small intestine

We investigated the distribution of FMRF amide-like immunoreactivity in the small intestine of the guinea pig. Immunoreactive nerve fibers were found mainly in the myenteric and submucous plexuses and in the inner circular muscle layer. The labeled processes contained variable proportions of small clear vesicles 30-40 nm in diameter and large granular vesicles 80-120 nm in diameter. The large granular vesicles showed heavy immunoreactivity. The antisera against FMRF amide crossreact with peptides belonging to the pancreatic polypeptide family; it has therefore been suggested that the FMRF amide immunoreactivity demonstrated in the small intestine is caused by a peptide that is biosynthetically related to, but not necessarily a member of, the pancreatic polypeptide family.     

Stretch-induced nerve conduction deficits in guinea pig ex vivo nerve

In the current communication, we characterized supraphysiologic elongations that elicited short-term nerve dysfunction. This was accomplished by assessing the electrophysiology of guinea pig tibial and peroneal nerves at predetermined elongation magnitudes. Results showed that a longitudinal supraphysiological stretch of lambda = 1.05 caused a 16% reduction in the mean compound action potential (CAP) amplitude. Upon relaxation to physiologic length, a full recovery in the CAP was observed. At lambda = 1.10, the CAP decreased by 50% with an 88% recovery after relaxation. For a supraphysiologic stretch of lambda = 1.20, severe conduction block with minimal acute recovery was observed. Latency also increased during periods of stretch and was proportional to the stretch magnitude. Additional studies showed some electrophysiological recovery during the sustained stretch phase. This attribute may be related to internal stress relaxation mechanisms. Since whole nerve elongations are averaged global deformations, we also used an incremental digital image correlation (DIC) technique to characterize the strain at the micro-tissue level. The DIC analysis revealed considerable heterogeneity in the planar strain field, with some regions exhibiting strains above the macroscale stretch. This non-uniformity in the strain map arises from structural inconsistencies of the nerve and we presume that zones of high local strain may translate into the observed conduction deficits.     

Ontogeny of mitochondrial steroidogenesis in the guinea pig gonad

To determine whether steroidogenesis in the developing guinea pig may be limited by the formation of pregnenolone, cholesterol side chain cleavage activity was ascertained at various stages of development. The conversion of [1,2-³H]cholesterol to [1,2-³H]pregnenolone was detected in mitochondria isolated from fetal guinea pig ovaries and testes as early as day 35 of gestation, while no metabolism was noted in day 30 animals. Moreover, no [l,2-³H]progesterone was formed during the 60 minute incubation. From day 35 of gestation to the day of birth, the percentage of pregnenolone formed per testis (total activity) increased, while total activity in the ovary declined. In contrast, gonadal mitochondria from adult guinea pigs converted cholesterol to both pregnenolone and progesterone and total activity in these animals was substantially higher than in their fetal counterparts. In the three females examined, the rate of pregnenolone and progesterone synthesis varied according to the stage of the estrous cycle during which these animals were sacrificed. Conversion of pregnenolone to progesterone was most rapid in the early luteal phase animal, while conversion of cholesterol to pregnenolone occurred more rapidly in the periovulatory animals than in ovarian mitochondria from the late luteal phase of the cycle. The results indicate that during prenatal and postnatal development of the gonad, cholesterol side chain cleavage activity changes and that mitochondria may acquire a Δ⁵-3β-hydroxysteroid dehydrogenase.     

Water transport in invertebrate peripheral nerve fibers

Osmotic and diffusion permeabilities (P_f and P_d) of invertebrate nerve fibers to tritiated water were measured to determine what water flux studies could reveal about "the nerve membrane" and to directly test the possibility of active transport of water into or out of invertebrate nerve fibers. P_f/P_d ratios for lobster walking leg nerve fibers were found to be about 20 ± 7 at 14°C. P_d measurements were made for squid giant axons at 25°C. and found to yield a value of 4 x 10^–4 cm.^–1 sec.^–1. When combined with the data of D. K. Hill for P_f, a P_f/P_d ratio of 21 ± 5 is obtained. These P_f/P_d ratios correspond to "effective pore radii" of about 16 ± 4 angstrom units, according to theories developed by Koefoed-Johnsen and Ussing and independently by Pappenheimer and his colleagues. Variations of water flux ratios with temperatures were studied and apparent activation energies calculated for both diffusion experiments and osmotic filtration experiments using the Arrhenius equation, and found to be close to 3 to 5 cal. per mole of water transferred. Cyanide (5 x 10^–3 molar) and iodoacetate (1 x 10^–3 molar) poisoned lobster leg nerve fibers showed no appreciable change in diffusion or osmotic filtration water effluxes. Caution in interpreting these proposed channels as simple pores was emphasized, but the possibility that such channels exist and are related to ionic flow is not incompatible with electrophysiological data.     

Citrate transport in guinea pig heart mitochondria.

Guinea pig heart mitochondria loaded with [-14C]citrate show exchanges of radioactivity at 30 degrees C with added citrate, L-malate and phosphoenolpyruvate. These exchanges are inhibited by benzene-1,2,3-tricarboxylate. Measurements of rates of citrate transport indicate that the activity of this transporting system is low in heart mitochondria compared to that observed in liver mitochondria. The K(m) values obtained indicate a similarity to those obtained in liver. Citrate oxidation by coupled mitochondria was also found to be slow at 30 degrees C but was inhibited by benzene-1,2,3-tricarboxylate. The role of mitochondrial citrate transport in control of glycolytic flux in the heart is discussed.     

Platelet-activating factor in the enteric nervous system of the guinea pig small intestine

Platelet-activating factor (PAF) is a proinflammatory mediator that may influence neuronal activity in the enteric nervous system (ENS). Electrophysiology, immunofluorescence, Western blot analysis, and RT-PCR were used to study the action of PAF and the expression of PAF receptor (PAFR) in the ENS. PAFR immunoreactivity (IR) was expressed by 6.9% of the neurons in the myenteric plexus and 14.5% of the neurons in the submucosal plexus in all segments of the guinea pig intestinal tract as determined by double staining with anti-human neuronal protein antibody. PAFR IR was found in 6.1% of the neurons with IR for calbindin, 35.8% of the neurons with IR for neuropeptide Y (NPY), 30.6% of the neurons with IR for choline acetyltransferase (ChAT), and 1.96% of the neurons with IR for vasoactive intestinal peptide (VIP) in the submucosal plexus. PAFR IR was also found in 1.5% of the neurons with IR for calbindin, 51.1% of the neurons with IR for NPY, and 32.9% of the neurons with IR for ChAT in the myenteric plexus. In the submucosal plexus, exposure to PAF (200-600 nM) evoked depolarizing responses (8.2 +/- 3.8 mV) in 12.4% of the neurons with S-type electrophysiological behavior and uniaxonal morphology and in 12.5% of the neurons with AH-type electrophysiological behavior and Dogiel II morphology, whereas in the myenteric preparations, depolarizing responses were elicited by a similar concentration of PAF in 9.5% of the neurons with S-type electrophysiological behavior and uniaxonal morphology and in 12.0% of the neurons with AH-type electrophysiological behavior and Dogiel II morphology. The results suggest that subgroups of secreto- and musculomotor neurons in the submucosal and myenteric plexuses express PAFR. Coexpression of PAFR IR with ChAT IR in the myenteric plexus and ChAT IR and VIP IR in the submucosal plexus suggests that PAF, after release in the inflamed bowel, might act to elevate the excitability of submucosal secretomotor and myenteric musculomotor neurons. Enhanced excitability of motor neurons might lead to a state of neurogenic secretory diarrhea.     

Coexistence and cooperation between neuropeptide Y and norepinephrine in nerve fibers of guinea pig vas deferens and seminal vesicle

Fluorescence immunocytochemistry of guinea pig vas deferens and seminal vesicle revealed dense networks of nerve fibers containing both neuropeptide Y (NPY) and dopamine-beta-hydroxylase (DBH), a marker for adrenergic neurons. The effects of norepinephrine (NE) and NPY on the smooth musculature of these organs were studied in vitro. NE inhibited the response to electrical nerve stimulation and increased the basic tension in the vas deferens and contracted the smooth muscle of the seminal vesicle, but had no effect on the contractile response to transmural stimulation in the latter organ. NPY had similar effects on the vas and vesicula, i.e. it inhibited the electrically induced contractions and had no effect on the basic tension. The results suggest a role for NPY as a transmitter that acts before the site of the neuromuscular junction to modulate the release of other transmitters from motor nerve fibers in the smooth musculature.     

Fine structure of fast-twitch and slow-twitch guinea pig muscle fibers

The guinea pig soleus muscle is a convenient model for the study of slow-twitch intermediate (STI) fiber ultrastructure because it is composed entirely of fibers of this class. Such fibers were compared with fast-twitch red (FTR) and fast-twitch white (FTW) fibers from the vastus lateralis muscle. FTW fibers are characterized by small, sparse mitochondria, a narrow Z line and, an extensive sarcoplasmic reticulum arranged primarily in longitudinal profiles at the A band and with numerous expansions at the I band. Abundant mitochondria with a dense matrix and subsarcolemmal and perinuclear aggregations are typical of FTR fibers. These fibers contain a plexus of sarcoplasmic reticulum at the A band and a less extensive network at the I band. The Z lines are wider (890 ± 74 Å) than those of FTW fibers (582 ± 62 Å). STI intermediate fibers are distinguished from other types by wide Z lines (1205 ± 58 Å), a faint M band, and a less extensive sarcoplasmic reticulum. Compared to FTR fibers, STI fiber mitochondria are usually smaller with less notable subsarcolemmal accumulations. FTW fibers have a more limited capillary supply, rarely contain lipid inclusions, and thus may be restricted to phasic activity. Extensive capillarity, mitochondrial and lipid context, and fast contraction times indicate possible phasic and tonic roles for FTR fibers. STI fibers, characterized by numerous lipid inclusions, extensive capillarity, relatively numerous mitochondria, but slow contraction-relaxation cycles, are morphologically suited for tonic muscle activity.     

Interleukin-1beta activates specific populations of enteric neurons and enteric glia in the guinea pig ileum and colon

Fos expression was used to assess whether the proinflammatory cytokine interleukin-1beta (IL-1beta) activated specific, chemically coded neuronal populations in isolated preparations of guinea pig ileum and colon. Whether the effects of IL-1beta were mediated through a prostaglandin pathway and whether IL-1beta induced the expression of cyclooxygenase (COX)-2 was also examined. Single- and double-labeling immunohistochemistry was used after treatment of isolated tissues with IL-1beta (0.1-10 ng/ml). IL-1beta induced Fos expression in enteric neurons and also in enteric glia in the ileum and colon. For enteric neurons, activation was concentration-dependent and sensitive to indomethacin, in both the myenteric and submucosal plexuses in both regions of the gut. The maximum proportion of activated neurons differed between the ileal (approximately 15%) and colonic (approximately 42%) myenteric and ileal (approximately 60%) and colonic (approximately 75%) submucosal plexuses. The majority of neurons activated in the myenteric plexus of the ileum expressed nitric oxide synthase (NOS) or enkephalin immunoreactivity. In the colon, activated myenteric neurons expressed NOS. In the submucosal plexus of both regions of the gut, the majority of activated neurons were vasoactive intestinal polypeptide (VIP) immunoreactive. After treatment with IL-1beta, COX-2 immunoreactivity was detected in the wall of the gut in both neurons and nonneuronal cells. In conclusion, we have found that the proinflammatory cytokine IL-1beta specifically activates certain neurochemically defined neural pathways and that these changes may lead to disturbances in motility observed in the inflamed bowel.     

Confocal laser microscopy of dystrophin localization in guinea pig skeletal muscle fibers

A confocal laser microscope was used to analyze the localization pattern of dystrophin along the sarcolemma in guinea pig skeletal muscle fibers. Hind leg muscles of the normal animals were freshly dissected and frozen for cryostat sections, which were then stained with a monoclonal antidystrophin antibody. In confocal laser microscopy, immunofluorescence staining in relatively thick sections could be sharply imaged in thin optical sections. When longitudinal and transverse sections of muscle fibers were examined, the immunostaining of dystrophin was seen as linearly aligned fluorescent dots or intermittent lines along the sarcolemma. In longitudinally cut muscle fibers, many fluorescent dots, but not all, corresponded to the sarcomere pattern, especially the I band. Sections cut tangential to the sarcolemma also showed a lattice-like pattern of longitudinal and transverse striations of fluorescent dots. Double staining for dystrophin and vinculin showed that the two proteins were not exactly colocalized. The end portions of muscle fibers were much more intensely stained with antidystrophin antibody than the central portions, following the contour of elaborate surface specializations at the myo-tendon junction. The staining pattern at the myo-tendon junction was also discontinuous. These confocal microscopic observations suggest that dystrophin may be localized in a nonuniform, discontinuous pattern along the sarcolemma and in some relationship with the underlying myofibrils.     

Immunohistochemical localization of calcitonin gene-related peptide and bombesin/gastrin-releasing peptide in nerve fibers of the rat,guinea pig and pig female genital organs

Summary The localization and distribution of calcitonin gene-related peptide (CGRP) and bombesin/gastrin-releasing peptide (GRP) immunoreactivity were studied in the rat, guinea pig and pig female genital organs with indirect immunohistochemical technique. In the rat, guinea pig and pig. CGRP and GRP immunoreactivities were localized in nerve fibers of the uterus, ovary and oviduct. Generally, CGRP-immunoreactive nerve fibers were intensely stained, while GRP-immunoreactive nerve fibers exhibited moderate immunoreactivity. The number of GRP-immunoreactive nerve fibers in these organs was lower in comparison with that of CGRP-immunoreactive nerve fibers. The pattern of distribution of these nerve fibers was very similar in different genital organs of all species studied. In the uterus of rat, guinea pig ang pig, CGRP-and GRP-immunoreactive nerve fibers and nerve bundles were observed in the muscular membrane and around blood vessels. Some delicate CGRP-and GRP-immunoreactive nerve fibers were also present in the submucous layer of the uterus. In the oviduct. CGRP-and GRP-immunoreactive nerve fibers were seen in the muscular membrane, around blood vessels and in the submucous layer. In the ovary, CGRP-and GRP-immunoreactive nerve fibers were distributed in medullary stroma, in close contact with blood vessels and between follicles of different stages of development.     

Potassium transport in dispersed mucosal cells from guinea pig stomach

Dispersed mucosal cells (approx. 70% parietal cells) prepared from guinea pig stomach maintained their cellular concentration of potassium (65--80 nmol potassium/10(6) cells) for at least 5 h in vitro. Uptake of 42K by dispersed gastric mucosal cells depended on temperature, H+ concentration and oxidative metabolism. Carbachol and, in some instances, gastrin caused a 40--50% increase in cellular uptake of 42K as a consequence of the ability of these agents to increase 42K influx. Ouabain reduced uptake of 42K by 70% but did not alter the effect of carbachol. Cellular uptake of 42K was not altered by histamine, prostaglandin, E1, glucagon, secretin, vasoactive intestinal peptide or C-terminal octapeptide of cholecystokinin. Uptake of 42K was also increased by dibutyryl cyclic AMP or dibutyryl cyclic GMP but not by cyclic AMP, cyclic GMP or their 8-bromo derivatives. Theophylline caused a small (10--15%) increase in 42K uptake and potentiated the increase caused by submaximal concentrations of carbachol. The increase in 42K uptake caused by either dibutyryl cyclic nucleotide and carbachol was additive.     

Immunohistochemical localization of calcitonin gene-related peptide and bombesin/gastrin-releasing peptide in nerve fibers of the rat, guinea pig and pig female genital organs

The localization and distribution of calcitonin gene-related peptide (CGRP) and bombesin/gastrin-releasing peptide (GRP) immunoreactivity were studied in the rat, guinea pig and pig female genital organs with indirect immunohistochemical technique. In the rat, guinea pig and pig, CGRP and GRP immunoreactivities were localized in nerve fibers of the uterus, ovary and oviduct. Generally, CGRP-immunoreactive nerve fibers were intensely stained, while GRP-immunoreactive nerve fibers exhibited moderate immunoreactivity. The number of GRP-immunoreactive nerve fibers in these organs was lower in comparison with that of CGRP-immunoreactive nerve fibers. The pattern of distribution of these nerve fibers was very similar in different genital organs of all species studied. In the uterus of rat, guinea pig and pig, CGRP- and GRP-immunoreactive nerve fibers and nerve bundles were observed in the muscular membrane and around blood vessels. Some delicate CGRP- and GRP-immunoreactive nerve fibers were also present in the submucous layer of the uterus. In the oviduct, CGRP- and GRP-immunoreactive nerve fibers were seen in the muscular membrane, around blood vessels and in the submucous layer. In the ovary, CGRP- and GRP-immunoreactive nerve fibers were distributed in medullary stroma, in close contact with blood vessels and between follicles of different stages of development.