An improved loose patch voltage clamp method using concentric pipettes

A method for noninvasive voltage-clamp recording from large cells is described. A firepolished pipette having two concentric barrels is pushed against the cell membrane, thereby electrically isolating a circular patch subdivided into an inner and an annular outer region. Both regions are held isopotential, but current is collected from the inner region only. The method electrically simulates a high resistance seal between pipette and cell membrane, allowing accurate and rapid voltage-clamp recording under conditions where the seal resistances actually obtained are low (near 1 M). This is useful in applications where one wishes to avoid enzymatic treatment.We provide details of electrode construction and voltage-clamp electronics, and present results obtained from frog skeletal muscle and leech neurons. For sodium channels of frog muscle, extensive data were previously obtained with other methods. There is good agreement between the earlier results and the measurements presented here.     

Preferential distribution of leukocytes in rat mesentery microvessel networks

Distribution of leukocytes in rat mesenteric microvessel networks was studied using intravital fluorescence video microscopy. A digital image analysis system was used to measure vessel diameters, flow velocities and leukocyte fluxes in 306 capillaries of 8 networks. Capillaries were defined as vessel segments connecting divergent to convergent branch points. Their topological position within the network was quantified by a generation number defined as the number of bifurcations between the capillary and the arteriole feeding the network.Proximal capillaries (generation numbers 4 and 5) were slightly but significantly smaller in diameter (8.9±0.4) m, mean ± SEM) than distal ones (generation numbers 20 and 21, 10.1±0.4 m). Average capillary flow velocity decreased markedly from 2.0±1.0 mm·s^–1 in proximal to 0.41 ±0.06 mm·s^–1 in distal capillaries. Average leukocyte concentration was 3.4±0.5·10⁹ l^–1 and thus significantly below systemic values (6.0·10⁹ l^–1) in proximal capillaries, and above in distal ones (11.7±2.6·10⁹l^–1).The analysis of flow and leukocyte flux partition at 138 bifurcations showed preferential distribution of leukocytes to the daughter capillary with higher flow rate. This suggests a tentative explanation for the observed leukocyte accumulation along the microvascular tree: due to their low fractional flow, proximal capillaries draw relatively leukocytepoor blood from the arteriole feeding the network; this leads to an increased leukocyte concentration in distal capillaries. As a consequence of the concomitant increase of capillary diameter with increasing generation number, leukocytes are preferentially flowing through larger capillaries and are excluded from small ones.     

Model experiments on the effect of bifurcations on capillary blood flow and oxygen transport

O₂-delivery by a single capillary is a function of the flow rate and thefraction of flow made up of red blood cells. Capillary flow rate in turn depends upon flow resistance which is determined by thefraction of capillary volume occupied by red blood cells. Experiments were carried out to study the relationship between these parameters in an in vitro model consisting of glass capillaries (I.D. 3.3–11.0 m) branching from a large bore feeding channel which was perfused at variable flow rates with suspensions of human red cells with different hematocrits. Capillary flow rates ranged from 0–10^–4 mm³ s^–1. The results indicate that the red cell flow fraction increases with increasing capillary flow rate and with decreasing feeding vessel flow rate. Capillary volume fraction occupied by red cells similarly depends on these two parameters, but is consistently lower than the red cell flow fraction. Capillary flow resistance increases with flow rate due to increasing volume fraction of cells. If the results obtained with the model system are applicable to in vivo capillaries it must be concluded that O₂-delivery by a single capillary is not linearly related to flow rate but increases more than proportionately with flow rate. Due to alteration of resistance with flow rate another type of autoregulation of capillary flow is proposed which tends to keep flow rate constant despite changes of driving pressure.Supported by the Deutsche Forschungsgemeinschaft     

The extracellular patch clamp: A method for resolving currents through individual open channels in biological membranes

The current contributions of individual ionic channels can be measured by electrically isolating a small patch of membrane. To do this, the tip of a small pipette is brought into close contact with an enzymatically cleaned membrane of a hypersensitive amphibian or mammalian muscle fiber. Current flowing through the pipette is measured. If the pipette contains cholinergic agonist at -molar concentrations, square pulse current waveforms can be observed which represent the activation of individual acetylcholine-receptor channels. The square pulses have amplitudes of 1 to 3 pA and durations of 10–100 ms.In order to obtain the necessary resolution, a delicate compromise had to be found between different experimental parameters. Pipettes with 1–3 m internal diameter and a steep final taper had to be used, extensive enzyme treatment was necessary, and conditions had to be found in which channels open at a relatively low frequency.     

Concentrative amino acid uptake at the serosal side of colon mucosa

4 healthy men had biopsies taken from the soleus and the lateral head of the gastrocnemius muscle for determination of capillary supply, fibre type distribution and fibre area.In serial transverse sections slow twitch (ST) and fast twitch (FT) fibres were identified histochemically by myofibrillar adenosine triphosphatase stains and capillaries visualized by a periodic acid Schiff stain after digestion of glycogen by -amylase.In soleus, ST and FT fibre type areas were approximately two times greater than in gastrocnemius. FT fibres (11060 m²) were 50% greater than ST fibres (7520 m²) in soleus, whereas no difference between FT and ST fibres was observed in gastrocnemius (4730 m² and 4310 m², respectively). Both muscles were mixed with respect to fibre types but in all subjects soleus had a higher relative content of ST fibres than gastrocnemius. The area occupied by ST fibres relative to total area was 64% in soleus and 50% in gastrocnemius.Capillary density was 288 capillaries·mm^–2 in soleus and 365 in gastrocnemius. Evaluated on the basis of mean number of capillaries in contact with fibres of each type relative to fibre type area (m^–2·10^–3) individual ST fibres had a richer capillary supply than FT fibres in both soleus (0.84 and 0.57, respectively) and gastrocnemius (1.12 and 0.92, respectively).Differences in organization of the capillary network between soleus and gastrocnemius were observed. Capillaries are running mainly parallel to the muscle fibres in gastrocnemius, but tortuously and/or frequently branching in soleus. Tortuously and/or frequently branching capillaries implies nutritional advantages, which will not be reflected in the quantitative data obtained from counts of capillaries made on transverse sectioned muscle. Therefore the absolute values given for soleus and gastrocnemius, respectively, cannot be directly compared.     

Ca2+ not cyclic AMP mediates the fluid secretory response to isoproterenol in the rat mandibular salivary gland: Whole-cell patch-clamp studies

We have performed whole-cell patch-clamp studies on dispersed seccretory cells of the rat mandibular gland to determine how -adrenergic stimulation causes fluid secretion. When the pipette contained a high K⁺ solution, the resting membrane potential averaged –33 mV±1.1 (SEM,n=34) and the clamped cell showed strong outward rectification. We monitored K⁺ and Cl^– currents for periods of 15 min by recording the currents needed to clamp the cell potential at 0 and –80 mV, respectively. Isoproterenol (1–2 mol/l) caused increases in the clamp current at 0 mV (the K⁺ current) and at –80 mV (the Cl^– current) in about 80% of cases, although the responses were variable in size and time-course; the responses were indistinguishable from those induced by acetylcholine or the Ca²⁺ ionophore, A23187. The -adrenergic antagonist, phentolamine (1–2 mol/l), had no effect on the response, but the -adrenergic antagonist, propranolol (10 mol/l), blocked it completely. The isoproterenol response could not be mimicked by application to either surface of the cell membrane, of cyclic AMP (100 mol/l), forskolin (1 or 20 mol/l) or cholera toxin (2.5 g/ml). However, increasing the Ca²⁺-chelating capacity of the pipette solution by raising its EGTA concentration from the customary 0.5 to 20 mmol/l, blocked the response to isoproterenol, suggesting that -adrenergic agonists activate Cl^– and K⁺ channels by raising cytosolic Ca²⁺. Since neomycin, which blocks phospholipase C, blocked the action of isoproterenol without impairing the cell responsiveness to A23187, it appears that isoproterenol, like muscarinic agonists, increased cytosolic Ca²⁺ via the phosphatidylinositol cycle.This project was supported by the National Health and Medical Research Council of Australia     

Electrophysiological analysis of sodium-transport in the colon of the frog (Rana esculenta)

Sodium transport and apical bioelectrical membrane properties were investigated in frog colonic epithelium in the absence and presence of the antidiuretic hormone arginine-vasotocin (AVT). Apical Na-permeability and intracellular Na-activity were evaluated by analysis of current-voltage relationships in the serosally K-depolarized tissue. Tissue- and apical membrane capacitance were measured by voltages step analysis. The frog colon was found to be a tight epithelium with a transepithelial resistance of 2.63±0.25 k·F (n=17). 85–90% of short circuit current (11.2±1.1 A·F·l^–1;n=17) was related to electrogenic Na-transport from mucosa to serosa. Graded doses of amiloride (<50 mol·l^–1) induced Michaelis-Menten-type inhibition kinetics. Serosal addition of 10^–6 mol·l^–1 AVT induced a significant increase in sodium current (25%), apical sodium permeability (19%) and tissue capacitance (4.3%) whereas intracellular Na-activity remained unchanged. There was a good correlation between increased Na-current and apical Na-permeability. No correlation was found between Na-current and membrane capacitance. Our results demonstrate that in contrast to other species the amphibian colon shows a natriferic reaction to AVT. We suggest that the regulation of Na-transport in frog colon is similar to that in the toad urinary bladder. It is caused by an activation of preexisting apical Na-channels and not by fusion of subapical cytoplasmic vesicles with the apical membrane.     

Influence of intratubular pressure on proximal tubular compliance and capillary diameter in the rat kidney

Tubular compliance is the response of tubular diameter to changes in intratubular pressure [7]. Proximal tubular compliance was determined directly by measurements of tubular diameter and pressure and indirectly using a mathematical model of tubular fluid flow based on measurements of the hydraulic pressure gradients along the tubule under free flow conditions and during an induced pressure reduction at the end of the proximal tubule. The two independent methods yielded similar values for compliance. Proximal tubular complicance was found to depend upon the intratubular pressure: tubular compliance was significantly higher (P<0.001) when the intratubular pressure was reduced below normal (1.0 m cm H₂O^–1) than when the pressure was increased above the control value (0.4 m cm H₂O^–1) Almost identical compliance values were measured in sodium pentobarbital and inactin anaesthetized rats (P>0.8).Intratubular pressure changes resulted in inverse changes in the diameters of the adjacent capillaries, suggesting that the peritubular capillaries are distensible structures.     

Preparation of freeze-dried cryosections for quantitative X-ray microanalysis of electrolytes in biological soft tissues

Summary A procedure is described which allows the evaluation of wet weight concentrations of diffusible substances in biological soft tissue on a cellular level by the use of energy dispersive X-ray microanalysis. Epithelia of frog skin and toad urinary bladder were used to prepare freeze-dried cryosections without the use of chemical fixatives, cryoprotectants, floating solutions or coating materials.The specimens were shock-frozen inl-propane (–180°C), cryosectioned (–80°C), sandwiched between collodion films and freeze-dried (–80°C). The analysis was performed in a scanning electron microscope at an acceleration voltage of 15 kV, probe current of 0.5 nA, using scanning areas of 1–2 m². The spatial resolution power using 1–2 m thick sections was about 0.7 m. In a superficial layer of about 30 m the analysis was found not to be influenced by tissue damage due to ice crystal formation. The mass loss during electron bombardement was restricted to constituents of the organic matrix (30%). No changes of characteristic radiation were observed for Na, K and Cl. Albumin standards, containing electrolyte concentrations in the range of biological interest, revealed linear calibration curves. To obtain reliable wet weight concentrations, the characteristic X-rays of the tissue were compared to those of an internal standard which was applied to the specimen prior to freezing and analysed simultaneously.Supported by grants from the Deutsche Forschungsgemeinschaft     

On the mechanism of β-adrenergic regulation of the Ca channel in the guinea-pig heart

Dose-response relations for the increase in the amplitude of Ca current (I _Ca) on external application of isoprenaline (ISP) and internally applied cyclic AMP (cAMP) or catalytic subunit of cAMP-dependent protein kinase (C subunit) were established in single ventricular cells of the guinea pig. An intracellular dialysis technique was used. The threshold concentration was for ISP 10^–9 M, for cAMP 3 M (pipette concentration to which 10^–5 M 3-isobutyl-1-methylxanthine was added) and for C subunit around 0.4 M (pipette concentration). The concentrations for the half-maximal effect were 3.7×10^–8 M (ISP), 5.0 M (cAMP) and 0.95 M (C subunit) and for the maximum effect 10^–6 M (ISP), 15–20 M (cAMP) and 3–4 M (C subunit). For all three agents the maximum increase in the Ca current density was similar (a factor of 3–4), suggesting that they converge on the same site of the Ca channel. Accordingly, the effects of cAMP and C subunit onI _Ca were non-additive to those of ISP. From these data the relationship both between concentrations of ISP and cAMP and between those of cAMP and active C subunit in terms of their effects onI _Ca could be estimated and were compared with those obtained in broken cell preparations.A competitive inhibitor of phosphorylation, 5-adenylyl-imidodiphosphate (5 mM), greatly reduced the effects of ISP and C subunit onI _Ca. Cell dialysis with 3 mM adenosine-5-(-thio)-triphosphate, which produces a dephosphorylationresistant phosphorylation, markedly potentiated the effects of ISP and cAMP onI _Ca.The results support the hypothesis that phosphorylation of a protein within, or close to, the Ca channel by cAMP-dependent protein kinase is the mechanism of -adrenergic stimulation.This work was supported by the Deutsche Forschungsgemeinschaft, SFB 38 (membranforschung), Projekt G, and H0579/6-2     

An effective transformation method for Hansenula polymorpha

Summary We have developed a rapid, simple transformation procedure for intact cells of Hansenula polymorpha. It is a modification of the LiAc method and can yield 10⁴–10⁵ transformants/g DNA. The use of stationaryphase cells, a high cell density per plate and a hear pulse at 50°C for 10 min are among major modifications of the original method. We have also found that, within the XhoI-SalI chromosomal fragment of Saccharomyces cerevisae containing the LEU2 gene, a sequence exists which supports autonomous replication of plasmid moleculles in H. polymorpha.     

Changes in diffraction patterns with length in single muscle fibres at rest

The addition of a simple X-Y sampling circuit to a closed circuit television system (CCTV) permits measurements of a narrow profile of the laser diffraction patterns from single frog muscle fibres at rest. Results confirm that maximum intensity occurs at 2.95–3.00 m, but a positive linear relationship between the dispersions of sarcomeres and sarcomere lengths from 2.7–3.8 m is obtained in four isolated single fibres.     

Somatic sodium channels of frog olfactory receptor neurones are inactivated at rest

Membrane excitability of acutely isolated olfactory receptor neurones (ORNs) of the grass frog (R. pipiens) was studied with the use of the whole-cell tight-seal patch recording technique. ORNs of the frog had a mean resting membrane potential of –52 mV, a mean input resistance of 1–2 G, and a mean capacitance of 4.5 pF. In the majority of cells examined (over 70%), short duration (several milliseconds) action potentials were elicited at the end of a hyperpolarising pulse (offspike) or following hyperpolarization of the membrane potential by injection of current. Under voltage-clamp conditions, a fast inward current followed by an outward current could be evoked upon depolarisation of the membrane. The fast inward current decayed with a time constant of 1–2 ms, with an e-fold decrease per 52 mV increase in voltage, and was blocked by the selective voltage-dependent sodium channel blocker tetrodotoxin (0.5–1 M). Steady-state inactivation studies revealed that the mean voltage for half-inactivation (V _1/2) was –82 mV (range –72 to –98 mV), which indicates that the voltage-dependent Na⁺ channels in the cell body or soma of frog ORNs are not available for conducting currents at the resting membrane potential. This finding raises the possibility that voltage-dependent Na⁺ channels may not play a significant role in sensory transduction at the soma. Our results indicate that ORNs of the frog are very efficient in transducing signals towards the brain since currents generated at the cilia will be directed towards depolarising the axon. The data also could account for the low number of single-cell action potentials encountered in extracellular recording experiments in vivo.     

Sensitivity to cyclic nucleotides and to Aspirin® of the kininogen-consuming system activated by adrenaline or carbamylcholine in rat blood

The activation by carbamylcholine of a kinin-forming system in rat blood in vivo or in vitro has been compared with that caused by adrenaline. After showing that free kinins were actually released into the circulation of the carbamylcholine-treated rat, experimentally induced changes in carbamylcholine or adrenaline activity were followed by measuring alterations in kinin precursor (kininogen) levels in plasma. Carbamylcholine was effective at 1.6–4.8 g/kg in the intact animal and optimal at 10^–8 M in oxalated rat blood in vitro. Aspirin^®, 10 mg/kg or 5×10^–4 M, inhibited the action of carbamylcholine and of adrenaline on kininogen. Salicylate or Indomethacin at similar levels were ineffective. N⁶-,2-O-dibutyryl-cyclic-3,5 adenosine monophosphate (diBu-cAMP) 2×10^–5 M, suppressed kininogen consumption in vitro; this inhibition could be overcome by 8-bromo-3–5-cyclic guanosine monophosphate (8-Br-cGMP), 10^–4 M. This compound potentiated the effect of adrenaline in the absence of diBu-cAMP. The inhibitory effect of Aspirin^® was decreased by 8-Br-cGMP. The action of autonomic effector drugs on kininogen consumption may be mediated by changes in the cAMP/cGMP ratio in circulatory target cells. The action of Aspirin^® may affect this system since it was partially overcome by 8-Br-cGMP. Since it was not reproduced by salicylate but could be mimicked by acetic anhydride, it most probably involves the transfer of the acetyl group of the drug to an as yet unknown acceptor cell protein. This moiety does not seem to be involved in prostaglandin-synthesis since Indomethacin, a very effective inhibitor of this process, failed to inhibit the action of either adrenaline or carbamylcholine on rat blood kininogen.     

Regulation of Na+ channels in frog lung epithelium: a target tissue for aldosterone action

Sodium transport across isolated lung tissue of the frog Xenopus laevis was measured in Ussing chambers under voltage-clamp conditions. Perfusing the lungs with NaCl-Ringer's solutions on both sides, a basal distinct amiloride-blockable Na⁺ current was present. Incubating the lungs with 1 mol/l aldosterone from the pleural side raised the short circuit current after a 1-h latent period. Maximal values were reached after 4–5 h of aldosterone treatment, at which time the transepithelial Na⁺ current was more than doubled compared to the control. The stimulatory effect was totally inhibited when the aldosterone treatment was preceded by incubation of the lung tissues with spironolactone in 2000-fold excess. In the presence of amiloride (0.5–8 mol/l) in the alveolar compartment, a Lorentzian noise component appeared in the power spectrum of the fluctuations in the short circuit current. This enabled the calculation of single Na⁺ channel current and Na⁺ channel density under both experimental conditions. Aldosterone stimulation did not change single Na⁺ channel current. On the other hand, the number of conducting Na⁺ channels increased in parallel with the transepithelial Na⁺ transport. This suggests that the alveolar epithelium may be a physiological target tissue for aldosterone. Since fluid absorption in the lung is secondary to active Na⁺ transport, aldosterone may be a potent regulator for maintaining the relatively fluid-free state of the lumen of the lung in some cases of fluid accumulation.     

Controlled Cell Deformation Produces Defined Areas of Contact between Cells and Ligand-Coated Surfaces

A method which allows precise control of the time of initiation and the area of contact of T cells with immobilized ligands has been developed. Cells are trapped in an asymmetric film that can be quantitatively thinned by reducing the film's capillary pressure. Ligands adsorbed to the base of the apparatus are forced into close contact with the cells as the air–liquid interface is drawn down. Using interference microscopy and microbeads to indicate the film height, the amount of thinning can be controlled to within 1 m. In this study, this system was used to produce contact areas of 182 and 356 m² between T cells and anti-CD3 coated surfaces. These contact areas were measured using fluorescent dye exclusion microscopy. This apparatus can be used for quantitative studies of T cell activation, as is reported in Patrick et al., J. Immunol. Method. 24:97–108, 2000. © 2001 Biomedical Engineering Society. PAC01: 8717-d, 8764Rr     

Characterization of the ATP-inhibited K+ current in canine coronary smooth muscle cells

Intracellular adenosine triphosphate (ATP)-inhibited K⁺ currents (I _{K, ATP} ) in canine coronary artery smooth muscle cells were characterized in the wholecell configuration using the suction pipette method. Cells dialysed internally with solutions containing 5 mM ATP (ATP_i) showed little detectable whole-cell current at potentials more negative than –30 mV. However, cells dialysed with ATP_i-free solutions developed a time- and voltage-independent current which reached a maximum of 132±25 pA at –40 mV about 10 min following patch rupture. After run-up, the current showed little run-down. Concentration-dependent inhibition by ATP_i yielded an inhibition constant (K _i of 350 M and a Hill coefficient of 2.3. In ATP_i-free solutions, the large current at –40 mV was reduced by glibenclamide with aK _i of 20 nM and a Hill coefficient of 0.95. Conversely, in 1 mM ATP_i solutions, the small current at –40 mV was increased by P-1075 from 8±2 pA to 143±33 pA, with a dissociation constant (K _d) of 0.16 M and a Hill coefficient of 1.7. The effect of P-1075 was antagonized by glibenclamide. Maximal current density elicited by either ATP_i depletion or external application of the channel opener P-1075 was similar with slope conductances of 81±10 pS/pF and 76±13 pS/pF respectively in the potential range of –90 to –40 mV. External Ca²⁺ had no effect on this current. Finally, in 1 mM ATP_i, 20 and 50 M adenosine increased the current slope conductance by 36±15% and 73±10% respectively between –90 to –40 mV. TheI _{K, ATP} although very small in these cells, was extremely effective in causing membrane potential hyperpolarization.     

Effects of shortening velocity and of oxygen consumption on efficiency of contraction in dog gastrocnemius

Previous observations have shown that, in isolated perfused dog gastrocnemii in situ, stimulated to aerobic rhythmic isotonic tetanic contractions (at about 40% of maximal isometric force), only about 20% of the overall metabolic power (proportional to the rate of O₂ consumption, O₂) was converted into mechanical power (Ẇ). Here we report that, in the same preparation, the maximal velocity during the shortening phase of each tetanus (v, mm s^–1) increased with the rate of energy dissipation, as given by the difference between O₂ and Ẇ (W kg^–1). The relationship between these variables was described by: v=2.85(O₂–Ẇ)^1.24 (R ²=0.85; n=17). A mathematical analysis of this equation shows that the overall mechanical efficiency (=ẆO₂ ^–1) decreased with increasing v (at constant O₂), whereas it increased with increasing O₂ (at constant v). The net effect of this state of affairs was that the decrease of over the entire range of work intensities was relatively minor (from 0.22 to 0.15), in spite of a large increase of v, (from 40 to 120 mm s^–1), thanks to the concomitant increase of O₂ (from 10 to 25 W kg^–1). So, under these experimental conditions, the energetics of work performance seems to be governed by two conflicting needs. The need for a sufficiently high shortening speed (and hence power output), itself requiring a sufficiently large energy dissipation rate, which, however, brings about a fall in . This is counteracted by the increased O₂, which in turn leads to an increased efficiency at the expense of a fall in shortening speed.This article was published in Human Muscular Function during Dynamic Exercise, Marconnet P, Saltin B, Komi P, Poortmans J (eds) Med Sport Sci 41 (series editors: M. Hebbelink, R.J. Shephard) pp. 1–9, Karger, Basel, 1996. It is reproduced with minor editorial modifications. Permission from Karger is gratefully acknowledged.     

Development of muscle-specific features in cultured frog embryonic skeletal myocytes

To study the development of muscle-specific features during myogenesis, we analysed the ultrastructure and voltage-dependent currents of frog embryonic skeletal myocytes maintained in culture for 10 days. The cells were maintained under culture conditions that prevented cell division, fusion and cell contacts with neuroblasts. The cell surface was estimated morphometrically and from cell capacity and the values obtained were used to calculate ion current densities. It was shown that the expression of all main types of voltage dependent ionic currents occurs during the first 3–5 days. Na⁺ maximum specific conductance at days 1–2 was low but by day 7 it showed a 20-fold increase. The magnitude of Na⁺ current densities increased 16-fold from day 1 (3.6 A/cm) to the day 7 (58.1 A/cm). The maximum specific K⁺ conductance increased almost 3-fold during the first 5 days. In contrast to the other types of currents, I_K undergoes qualitative changes. Sodium action potentials, whose amplitude and time course depend on g_Na/g_K ratio, appeared from day 4 in culture, when myofibrils and the T-system also developed. The amplitude of DHP-sensitive slow I_Ca increased in parallel with the development of the T-membrane. I_Ca,S density per unit of T-membrane area reached an equilibrium of ca., 17 A/cm² on the day 4 and then remained stable until the end of the period of observation. These studies demonstrate that muscle-specific characteristics including morphology and excitatory properties begin to develop on the third day and resemble those of adult muscle cells by the sixth day in culture.     

Conduction of nerve impulses within the pineal system of frog

Summary The conduction of nerve impulses within the pineal system of the frog has been investigated. Uninterrupted transmission of nerve impulses was seen after electrical stimulation of the intracranial pineal tract and recording of action potentials from the extracranial portion of the pineal nerve. The conduction velocities were at 2–9 m/sec (medullated fibres of 3–8 diameter), 1–2 m/sec (intermediate group, 1–3 ) and 0.2–0.9 m/sec (unmyelinated fibres, 0.2–0.8 ). The highest speed of conduction was observed in R. catesbyana where the slowest fibres were conducting at velocities of the same range as the fastest nerve fibres in some specimens of R. temporaria and R. esculenta. It was concluded that the conduction of afferent impulses from the frontal organ to the posterior commissure of the midbrain is based mainly on unmedullated fibres.This work was supported by grants from the Deutsche Forschungsgemeinschaft toward the cost of apparatus.A preliminary account of this work was presented at the Spring Meeting of the Deutsche Physiologische Gesellschaft at Würzburg, April 1966.