FISHERY BULLETIN: VOL. 71. NO. 4 



between growth bands while KOH removed 

 adherent flesh. Staining techniques of Hollister 

 (1934), Galtsoff (1952), Daiber (1960), and La 

 Marca (1966) were not effective in staining 

 growth marks differentially. Puffer vertebrae 

 have deeply concave centra and had to be sliced 

 lengthwise in order to measure straight-line 

 distances. Vertebrae No. 4 and No. 5 with the 

 neural spines removed were placed dorsal sur- 

 face up in a V-shaped slit in a wood block and 

 were sliced with a hand-held razor blade. All 

 vertebral measurements were made on vertebra 

 No. 4 using an ocular micrometer in a dissect- 

 ing microscope at 15 x under reflected light. 

 The marginal width or distance between the 

 last annulus and the centrum edge was mea- 

 sured on the posterior segment of vertebra No. 4. 



AGE DETERMINATION 



Vertebrae and saccular otoliths exhibited 

 growth marks, although only the growth marks 

 on vertebral centra were both distinct and con- 

 sistent. Growth marks on otoliths were ambigu- 

 ous and could not be distinguished in larger 

 fish. Cleithra, opercles, jaw bones, and fin rays 

 exhibited no discernible growth marks. 



Growth marks on northern puffer vertebrae 

 formed distinct steps on the centrum surface 

 which are best seen when a vertebra is rested 

 on one end at a 45° angle from the vertical 

 and is viewed looking down on the centrum face. 

 The step is more opaque and its surface texture 

 appears coarser under magnification in reflect- 

 ed light than the rest of the centrum. A narrow, 

 dark, translucent ring appears along the inner 

 edge of each step. The step and narrow dark 

 band form a continuous ring on the centrum 

 and constitute the annulus. Broad opaque 

 bands with uniform surface texture lie between 

 the narrower annuli. Together these features 

 represent 1 yr's growth. Season marks on 

 black bullhead, Ictalurus melas, vertebrae are 

 narrow, translucent bands associated with a 

 depression in the centrum surface and alternat- 

 ing with broad, opaque bands (Lewis, 1949). 

 In contrast, narrow dark annual bands on chan- 

 nel catfish, /. punctatus, were usually associat- 

 ed with a ridge (Appelget and Smith, 1951). 



956 



Five age groups of northern puffer were 

 found by considering the marks on vertebrae 

 to be annuli. 



There was excellent agreement among verte- 

 brae No. 3-No. 7 in the number and position of 

 annuli. Age could not be assigned because of 

 disagreement in less than 1% of the fish. How- 

 ever, age could not be determined in 7% of the 

 fish because of false annuli, crowding of an- 

 nuli at the centrum edge, and changes in the 

 surface texture of centra in older fish. False 

 annuli were faint dark bands which were not 

 associated with a distinct step. Another ex- 

 traneous mark was a shallow depression form- 

 ing a continuous ring on the centrum which 

 was not a definitive step. This feature was 

 found on black bullhead vertebrae (Lewis, 1949). 

 False annuli were most frequent in the first 

 growth zone (Figure la). With increasing age, 

 annuli became crowded at the centrum edge 

 (Figure lb), and the surface texture of centra 

 became more coarse and irregular, especially 

 near the edge (Figure Ic). However, there was 

 no indication that annuli at the centrum edge 

 were obscured by crowding in the 58 age III 

 and 17 age IV fish examined. 



VALIDATION OF THE ANNULUS 



The annular step on vertebrae of the northern 

 puffer met four of Hile's (1941) criteria for 

 validating the annulus as a true year mark. 

 First, a single annulus was formed each year. 

 No annuli were present on vertebrae of year- 

 lings in April and early May, but a distinct r 

 ring became visible at the outer edge of the 

 centrum late in May when growth resumedf 

 (Figure 2). Annulus formation began in May.j 

 and by August most yearlings had formed an' 

 annulus (Figure 3). The fish taken in April 

 with a single annulus and wide marginal width 



Figure 1. — Vertebrae of the northern puffer: (a) verte- 

 brae from a 2-yr-old northern puffer showing a false 

 annulus adjacent to the first annulus, (b) vertebrae from a 

 5-yr-old northern puffer on which the third annulus 

 appears double because of the expanded step, (c) vertebrae 

 from a 3-yr-old northern puffer, (d) expanded annular 

 step on the vertebrae from a 1-yr-old northern puffer. 



I 



