15.2. 1975

Specialia

Zusammen/assung. E s k o n n t e gezeigt w e r d e n , dass elektrische S t i m u l a t i o n des l a t e r a l e n H y p o t h a l a m u s m i t einer V e r z 6 g e r u n g y o n 2-6 m i n F r e s s e n ausl6st. Selbstr e i z v e r s u c h e zeigten, dass diese S t i m u l a t i o n s u c h belohn e n d wirkt. ~ h n l i c h v e r z 6 g e r t e s F r e s s e n k a n n s u c h d u r c h

~ Research supported by the Swiss National Science Foundation Grant No. 3.8790.72.

189

R e i z u n g oder s p r e a d i n g d e p r e s s i o n in a n d e r e n H i r n g e b i e t e n ausgel6st w e r d e n . E s w i r d v e r m n t e t , dass alle diese Ph~tnomene auI e i n e m g e m e i n s a m e n M e c h a n i s m u s basieren. J. P. HUSTON, K. ORNSTEIN a n d B. SIEGFRIED 19

I~r o/ Pharmacology, Ueaiversity o/ Z~rich, Gloriastrasse 32, CH-8006 Zi~rich (Switzerland), 3 October 1974.

Cuticular Gas in Marine Decapod Crustacea A film of gas, p e r h a p s n o m o r e t h a n a few m i c r o n s thick, on t h e b o d y surface h a s b e e n s u g g e s t e d as t h e basis of t h e p r e s s u r e s e n s i t i v i t y s h o w n b y c e r t a i n m a r i n e c r u s t a c e a a, b u t u n t i l r e c e n t l y it h a s n o t b e e n possible t o e s t a b l i s h t h e e x i s t e n c e of such m i n u t e v o l u m e s ot gas e x p e r i m e n t a l l y . T h e ultrasonic t e c h n i q u e s c u r r e n t l y available are e i t h e r non-specific for gas 2 or d e s i g n e d t o d e t e c t m o v i n g b u b b l e s using t h e D o p p l e r shift principle a, 4. A n a l t e r n a t i v e m e t h o d s,6, b a s e d on m e a s u r e m e n t of t h e h a r m o n i c w a v e f o r m d i s t o r t i o n caused b y gas in a n a c o u s t i c field h a s b e e n d e v e l o p e d in t h e D e p a r t m e n t oi E l e c t r o n i c a n d E l e c t r i c a l E n g i n e e r i n g a t t h e U n i v e r s i t y of B i r m i n g h a m . This t e c h n i q u e , w h i c h has t h e a d v a n t a g e of b e i n g specific for gas, is t h u s s u i t a b l e for t h e d e t e c t i o n a n d e s t i m a t i o n of small, s t a t i o n a r y gas v o l u m e s in a liquid m e d i u m , a n d h a s b e e n used h e r e for t h e d e t e c t i o n of gas o n t h e b o d y surface of d e c a p o d e r u s t a c e a . Materials and meIhod. E x p e r i m e n t s were carried o u t w i t h t h e t r a n s m i t a n d receive acoustic t r a n s d u c e r s a r r a n g e d a b o u t 5-7 m m a p a r t (Figure la) a n d i m m e r s e d in k e r o s e n e to a v o i d electrolysis a t t h e metalIic surfaces of t h e t r a n s d u c e r s . A n a c o u s t i c a l l y ' t r a n s p a r e n t ' t e s t cell,

c o n t a i n i n g w a t e r boiled at r e d u c e d p r e s s u r e to r e m o v e a n y r e s i d u a l air bubbles, was p o s i t i o n e d b e t w e e n t h e t w o transducers and the apparatus calibrated by generating a k n o w n v o l u m e of gas electrolytically on t h e end of a fine c o p p e r wire, i n s u l a t e d to t h e t i p a n d held in t h e acoustic field. Legs r e m o v e d f r o m living or freshly killed s p e c i m e n s of t w o p r e s s u r e - s e n s i t i v e species of crab, Carcimas maenas 7 a n d Macropipus holsatus s, were m o u n t e d in a r u b b e r

1 p. S. B. DmBY, Nature, Lond. 191, 366 (1961). 2 D. N. WALDER,A. EVANS and H. V. HEMPLEMAN,Lancet 1, 897 (1968). a M. GILLIS, P. L. PETERSONand M. T. t{ARAGIANES,Nature, Lond. 217, 965 (1968). A. EvAns, E. E. P. BARNARDand D. N. WALDER,Aerospace Med. 43, 1095 (1972). 5 V. G. WELSBY, Proe. Joint Conf. Industrial Ultrasonics, Inst. ~Eleetronic and Radio EngTs. Conf, Prom 16, 97 (1969). s V. G. W~LSB'Z and M. H. SAFAR,Acustica 22, 177 (1969170). E. NA'ZLORand R. J. A. ATKINSOG Symp. Soc. exp. Biol. Med. 26, 395 (1972). a E. MORGAY,Crustaceana 13, 275 (1967).

Glass rod

a)

It-

Transmit

transducer

Acoustically b) transparent

membrane [eceive

transducer

Test specimen

Water boiled under reduced

Paraffin bath

pressure

Fig. 1. a) Diagram of the apparatus. The transmit transducer was made from paired lead-zirconate plates mounted on aluminium alloy blocks and enclosed in an Muminium alloy housing while the receiver contained a single plate embedded ila araldite in an atuminium alloy block. Kerosene was preferred to water as an acoustic coupling fluid to avoid generating gas electrolytieaUy on the metallic surface of the transducers. The specimen under investigation was lowered into a test ceil with acoustically-transparent membrane walls, and containing water boiled under reduced pressure to remove air bubbles. The fundamental and harmonic waveforms were displayed on an oscilloscope and the harmonic distortion component amplified and rectified for monitoring on a pen recorder when required, b) Diffusion cell with dialysis membrane collar.

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sleeve on t h e end of a glass rod a n d t h e possible e x i s t e n c e of gas on t h e surface i n v e s t i g a t e d b y lowering t h e leg into t h e cell b e t w e e n t h e t r a n s d u c e r s . ]?esults and discussion. The resulting acoustic h a r m o n i c d i s t o r t i o n (Figure 2) i n d i c a t e d gas w h i c h r a n g e d in v o l u m e f r o m 0.3 • 10 -6 t o 1.3 • 10 -6 ml in t h e d i f f e r e n t e x p e r i m e n t s . A s s u m i n g an even d i s t r i b u t i o n , t h e g r e a t e s t v o l u m e of gas r e c o r d e d would c o r r e s p o n d to a layer no m o r e t h a n 40 n m t h i c k a n d it s e e m s m o r e p r o b a b l e t h a t t h e gas is d i s t r i b u t e d in a mosaic of small pockets. H o w e v e r , it is d o u b t f u l w h e t h e r t h e gas r e c o r d e d here was g e n e r a t e d as t h e result of a n electric p o t e n t i a l difference across t h e cuticle, as suggested b y DIGB'Z 9. Similar v o l u m e s h a v e b e e n d e t e c t e d on sections of c a r a p a c e r e m o v e d f r o m c r a b s f o u n d d e a d on t h e b o t t o m of t h e h o l d i n g t a n k s in t h e Zoology D e p a r t m e n t a q u a r i u m , a n d on e l y t r a r e m o v e d f r o m d e a d a n d living cockroaches. The acoustic p r e s s u r e c h a n g e s g e n e r a t e d b e t w e e n t h e t r a n s d u c e r s were small, a b o u t 10 millibars (lkNm -2) p e a k - t o - p e a k , a n d unlikely to effect c o m p r e s s i o n of t h e v e i n s in t h e elytra, a n d it t h e r e f o r e seems p r o b a b l e t h a t t h e gas d e t e c t e d in t h e s e e x p e r i m e n t s h a d s i m p l y ads o r b e d on t o t h e surface of t h e i n t e g u m e n t . I n a f u r t h e r series of e x p e r i m e n t s legs were a r n p u t a t e d f r o m Carcinus a n d t r a n s f e r r e d to t h e acoustic c h a m b e r u n d e r w a t e r . P o t e n t i a l differences of 35 50 mV, inside negative, h a d b e e n p r e v i o u s l y r e c o r d e d across t h e integum e n t of t h e leg, a n d gas could c o n c e i v a b l y originate electrolytically u n d e r t h e s e conditions. O n l y on one occasion was gas d e t e c t e d a n d t h e n in m u c h smaller q u a n t i t i e s t h a n above. The a m p u t e e h a d b e e n k e p t s u b m e r g e d for a b o u t a w e e k p r i o r t o t h e e x p e r i m e n t , b u t h a d b e e n e x p o s e d p r e v i o u s l y in t r a n s i t , a n d it is possible t h a t m i n u t e q u a n t i t i e s of air could h a v e s u r v i v e d over t h i s period in m i c r o c a v i t i e s in t h e cuticle ~0 A l t h o u g h t h e periodic e x p o s u r e of littoral a n i m a l s could t h u s confer u p o n t h e m a m e c h a n i s m of p r e s s u r e t r a n s d u c tion, it would a p p e a r unlikely t h a t such a n i m p o r t a n t e n v i r o n m e n t a l v a r i a b l e should be p e r c e i v e d so fortuitously. M a n y p r e s s u r e - s e n s i t i v e a n i m a l s are plank-t o n i c ~-~a a n d are n o t n o r m a l l y e x p o s e d t o t h e air. DIGBY p o s t u l a t e d a n electrogenesis of gas a t t h e b o d y surface on t h e basis of a series of e x p e r i m e n t s in w h i c h he d e m o n s t r a t e d h o w t h e p o t e n t i a l difference r e c o r d e d

across t h e i n t e g u m e n t of t h e p r a w n Palaemonetes varians, a n d b e t w e e n diffusion cells of d i f f e r e n t ionic c o n c e n t r a t i o n s v a r i e d w i t h c h a n g e s in a m b i e n t pressure. These e x p e r i m e n t s are r e a d i l y r e p e a t e d . U s i n g c o p p e r electrodes (cf DIGBu p r e s s u r e - i n d u c e d c h a n g e s in p o t e n t i a l h a v e b e e n r e c o r d e d f r o m isolated legs of Carcinus, stage 2 larvae of t h e l o b s t e r Homarus, a n d also f r o m a c o m p a rable diffusion cell 1,. These results are c o n s i s t e n t w i t h t h e c o m p r e s s i o n of a gas p h a s e b u t it is difficult t o a s c e r t a i n w h e t h e r this is l o c a t e d a t t h e surface of t h e i n t e g u m e n t (or diffusion cell m e m b r a n e ) or on t h e surface of t h e metallic electrode itself. W e h a v e a t t e m p t e d t o locate t h e gas in a m o d e l cell s y s t e m in w h i c h sea w a t e r was separ a t e d f r o m slightly saline t a p w a t e r b y a collar of dialysis m e m b r a n e (Figure lb). The p o t e n t i a l difference d e v e l o p e d across t h e m e m b r a n e was f o u n d to be 70 80 mV, inside negative, a n d a f u r t h e r --15 m V was r e c o r d e d a t t h e j u n c t i o n b e t w e e n t h e c o p p e r wire c a t h o d e a n d t h e sea w a t e r w i t h i n t h e cell. Overall p o t e n t i a l c h a n g e s of up t o 20% h a d b e e n p r e v i o u s l y r e c o r d e d f r o m similar cells following an increase of p r e s s u r e of up t o 1250 m b (125 knm-2). I n our e x p e r i m e n t s t h e cell was p o s i t i o n e d w i t h t h e m e m b r a n e collar in t h e c e n t r e of t h e acoustic field a n d a w e a k c u r r e n t p a s s e d b e t w e e n t h e electrodes. Gas w a s d e t e c t e d o n l y w h e n t h e t i p of t h e c o p p e r c a t h o d e was l o c a t e d w i t h i n t h e acoustic field. No gas was r e c o r d e d w h e n t h e electrode was p a r t i a l l y w i t h d r a w n f r o m t h e cell t o a p o s i t i o n o u t s i d e t h e acoustic field, s u g g e s t i n g t h a t in t h i s e x p e r i m e n t t h e gas was g e n e r a t e d p r i m a r i l y a t t h e surface of t h e m e t a l l i c electrode. H o w e v e r , as DIGBY I5 has m o r e r e c e n t l y p o i n t e d out, if t h e m e m b r a n e is too p o r o u s a n y gas p r o d u c e d a t its surface w o u l d be r e m o v e d b y e l e c t r o - o s m o t i c effects. C r u s t a c e a n cuticle is m u c h less porous, a n d has b e e n s h o w n t o f u n c t i o n as a n i n t e r m e d i a t e electrode a t w h i c h h y d r o g e n is g e n e r a t e d w h e n a c u r r e n t is p a s s e d across it, b u t t h e e v i d e n c e t h a t gas is g e n e r a t e d electrolytically a t t h e b o d y surface in t h e n a t u r a l s i t u a t i o n a p p e a r s inconclusive.

Rdsumd. On a d6cel6 la pr6sence d ' u n e t o u c h e de gaz d'/~peupr~s 40 n m d ' @ a i s s e u r sur la cuticule de c e r t a i n s crustac6s d6capodes m a t i n s en e n r e g i s t r a n t la d i s t o r s i o n h a r m o n i q u e qua ce gaz a caus6e l o r s q u ' o n le s o u m e t 5~ u n c h a m p acoustique. Q u o i q u ' o n s i t d 6 m o n t r 6 qua le gaz p o u r r a i t 6tre adsorb6 sur la surface, une origine 61ectrol y t i q u e est ~ la limite du possible. E. MORGAN16 a n d V. G. WELSBY

Department o/ Zoology and Comparative Physiology, and Department o/Electronic and Electrical Engineering, University o/ Birmingham, P.O. Box 363, Edgbaston, Birmingham B I 5 2 T T (England), 73 August 1974.

J--J2 sac Fig. 2. Trace obtained on lowering the 3rd walking leg of the shore crab Carcinus maenas into, and subsequently removing it from, the acoustic field. The leg was positioned manually and the harmonic distortion caused by gas was found to be proportional to the amount of leg located between the transducers, suggesting a uniform distribution over the surface of the integument. The volume of gas involved is indicated by the extent of the vertical deflection,

9 p. S. B. DIGBY, Symp. Zool. Soc., Loud. 19, 159 (1962). lo M. STRASBeRG,J. aeoust. Soe. Am. 31, 163 (1959). 11 A. C. HARDYand R. BAINBRII)G~, Nature, Lond. 767, 354 (1951). ,,z E . W . KNIGHT-JONES and S. Z. QASlM, Nature, Lond. 175, 941 (1955). la E. R. 13AYLORand F. E. SMITH, ill Recent. Advances Invertebrate Pyhsiology Univ. of Oregon Press, Eugene 1957), p. 21. 14 ]~. MORGAN,

unpublished.

~a P. S. t3. DIGBY, Symp. Soe. exp. Biol. Mad. 26, 445 (1972). 16 Part of this work was carried out by one of us (E.M.) while at the Marine Biological Station, Port Erin. It is a pleasure to recognize the hospitality of the Director and his staff.